MedWorm.com provides a medical RSS filtering service. Over 6000 RSS medical sources are combined and output via different filters. This feed contains the latest items from the 'BioTechniques' source. Fri, 03 Feb 2012 18:24:30 +0100 http://www.medworm.com/index.php?rid=5597530&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229718%26dopt%3DAbstract Authors: Blow NS PMID: 22229718 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597530 Sun, 01 Jan 2012 05:00:00 +0100 5597530 http://www.medworm.com/index.php?rid=5597529&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229719%26dopt%3DAbstract Authors: Lo PC, Nybo K PMID: 22229719 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597529 Sun, 01 Jan 2012 05:00:00 +0100 5597529 http://www.medworm.com/index.php?rid=5597528&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229720%26dopt%3DAbstract Authors: Dorman N PMID: 22229720 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597528 Sun, 01 Jan 2012 05:00:00 +0100 5597528 http://www.medworm.com/index.php?rid=5597527&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229721%26dopt%3DAbstract Authors: Nybo K PMID: 22229721 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597527 Sun, 01 Jan 2012 05:00:00 +0100 5597527 http://www.medworm.com/index.php?rid=5597526&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229722%26dopt%3DAbstract Authors: Webb S PMID: 22229722 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597526 Sun, 01 Jan 2012 05:00:00 +0100 5597526 http://www.medworm.com/index.php?rid=5597525&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229723%26dopt%3DAbstract Authors: Nybo K PMID: 22229723 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597525 Sun, 01 Jan 2012 05:00:00 +0100 5597525 http://www.medworm.com/index.php?rid=5597524&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229724%26dopt%3DAbstract In this study, we evaluated the potential of the wheat-germ cell-free translation system for producing human HtrA3. HtrA3 underwent autocleavage when synthesized at 17°C. When the synthesis temperature was lowered to 4°C, full-length HtrA3 was successfully produced and proteolytically active. Catalytic site serine substitution with alanine (S305A) stabilized HtrA3 while abolishing its protease activity. This mutant was readily synthesized and stable at 17°C. When used with glutathione S-transferase (GST) pull-down assay, S305A HtrA3 was a valuable bait in searching for endogenous HtrA3 binding proteins. Thus, we demonstrated the unique utility of the wheat-germ cell-free translation system for producing and characterizing human HtrA3. These strategies will be likely applicable to a wide... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597524 Sun, 01 Jan 2012 05:00:00 +0100 5597524 http://www.medworm.com/index.php?rid=5597523&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229725%26dopt%3DAbstract In this study, we have developed a robust cryohistological method that allows imaging of virtually any type of plant cell or tissue while preserving fluorescent protein signals and maintaining excellent cellular and subcellular morphology. This method involves modified fixation of plant tissues (i.e., leaves, stems, and petioles), infiltration in a sucrose gradient, freezing, and collection of cryosections directly onto a cryoadhesive tape. Using this method followed by microscopic analysis, we demonstrated a localized accumulation of green fluorescent protein (GFP) in Nicotiana benthamiana plants agroinfiltrated with the movement-incompetent tobacco mosaic virus-based vector and systemic accumulation of GFP in plants infiltrated with the movement-competent vector. Overall, this simple cry... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597523 Sun, 01 Jan 2012 05:00:00 +0100 5597523 http://www.medworm.com/index.php?rid=5597522&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229726%26dopt%3DAbstract We describe the application of a simple and low cost dielectrophoretic device for picking out and relocating single target cells. The device consists of a single metal electrode and an AC signal generator. It does not require microfabrication technologies or sophisticated electronics. The dielectrophoretic manipulator also discriminates between live and dead cells and is capable of redistributing intracellular organelles. PMID: 22229726 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597522 Sun, 01 Jan 2012 05:00:00 +0100 5597522 http://www.medworm.com/index.php?rid=5597521&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229727%26dopt%3DAbstract Authors: Ohashi K, Kiuchi T, Shoji K, Sampei K, Mizuno K Abstract The bimolecular fluorescence complementation (BiFC) assay is a method for visualizing protein-protein interactions in living cells. To visualize the cofilin-actin interaction in living cells, a series of combinations of the N- and C-terminal fragments of Venus fused upstream or downstream of cofilin and actin were screened systematically. A new pair of split Venus fragments, Venus (1-210) fused upstream of cofilin and Venus (210-238) fused downstream of actin, was the most effective combination for visualizing the specific interaction between cofilin and actin in living cells. This pair of Venus fragments was also effective for detecting the active Ras-dependent interaction between H-Ras and Raf1 and the Ca(2+)-depen... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597521 Sun, 01 Jan 2012 05:00:00 +0100 5597521 http://www.medworm.com/index.php?rid=5597520&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22229728%26dopt%3DAbstract Authors: Berlec A, Strukelj B, Strukelj B Abstract The growing popularity of the lactic acid bacterium Lactococcus lactis has increased demand for novel high-throughput cloning methods. Here we describe a general TA-cloning methodology and demonstrate its feasibility using the plasmid pNZ8148. PCR products were directly ligated into a linear, PCR-amplified and XcmI-digested pNZ8148 derivative that was termed pNZ-T. Cloning using pNZ-T yielded a high proportion of insert-containing plasmids on transformation. Although demonstrated with L. lactis, the technique presented here is organism-independent and can be implemented in other plasmids. PMID: 22229728 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5597520 Sun, 01 Jan 2012 05:00:00 +0100 5597520 http://www.medworm.com/index.php?rid=5531231&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150324%26dopt%3DAbstract Authors: Blow NS PMID: 22150324 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531231 Thu, 01 Dec 2011 05:00:00 +0100 5531231 http://www.medworm.com/index.php?rid=5531230&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150325%26dopt%3DAbstract Authors: Lo PC, Nybo K PMID: 22150325 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531230 Thu, 01 Dec 2011 05:00:00 +0100 5531230 http://www.medworm.com/index.php?rid=5531229&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150326%26dopt%3DAbstract Authors: Dorman N PMID: 22150326 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531229 Thu, 01 Dec 2011 05:00:00 +0100 5531229 http://www.medworm.com/index.php?rid=5531228&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150327%26dopt%3DAbstract Authors: Nybo K PMID: 22150327 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531228 Thu, 01 Dec 2011 05:00:00 +0100 5531228 http://www.medworm.com/index.php?rid=5531227&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150328%26dopt%3DAbstract Authors: Webb S PMID: 22150328 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531227 Thu, 01 Dec 2011 05:00:00 +0100 5531227 http://www.medworm.com/index.php?rid=5531226&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150329%26dopt%3DAbstract Authors: PMID: 22150329 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531226 Thu, 01 Dec 2011 05:00:00 +0100 5531226 http://www.medworm.com/index.php?rid=5531225&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150330%26dopt%3DAbstract Authors: Nybo K PMID: 22150330 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531225 Thu, 01 Dec 2011 05:00:00 +0100 5531225 http://www.medworm.com/index.php?rid=5531224&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150331%26dopt%3DAbstract Authors: O'Callaghan N, Baack N, Sharif R, Fenech M Abstract Telomere shortening is an important risk factor for cancer and accelerated aging. However, it is becoming evident that oxidatively damaged DNA within the telomere sequence may also cause telomere dysfunction. Here we describe a reliable, cost-effective quantitative PCR (qPCR)-based method to measure the amount of oxidized residues within telomeric DNA that are recognized and excised by formamidopyridine DNA glycosylase (FPG). We also report that in an in vitro model of oxidative stress oxidized base lesions measured using this method are more prevalent within telomeric sequences. Furthermore, this method is sufficiently sensitive to detect changes in oxidative stress induced by zinc deficiency and hydrogen peroxide within... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531224 Thu, 01 Dec 2011 05:00:00 +0100 5531224 http://www.medworm.com/index.php?rid=5531220&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150332%26dopt%3DAbstract Authors: Hoon S, Zhou B, Janda KD, Brenner S, Scolnick J Abstract Traditional methods for selecting aptamers require multiple rounds of selection and optimization in order to identify aptamers that bind with high affinity to their targets. Here we describe an assay that requires only one round of positive selection followed by high-throughput DNA sequencing and informatic analysis in order to select high-affinity aptamers. The assay is flexible, requires less hands on time, and can be used by laboratories with minimal expertise in aptamer biology to quickly select high-affinity aptamers to a target of interest. This assay has been utilized to successfully identify aptamers that bind to thrombin with dissociation constants in the nanomolar range. PMID: 22150332 [PubMed - in proc... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531220 Thu, 01 Dec 2011 05:00:00 +0100 5531220 http://www.medworm.com/index.php?rid=5531198&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150333%26dopt%3DAbstract Authors: Grupillo M, Lakomy R, Geng X, Styche A, Rudert WA, Trucco M, Fan Y Abstract Intracellular staining is a widely used flow cytometry (FCM)-based technique to detect the expression of cytoslio nucleic antigens. However, intracellular staining of cells expressing cytosolic fluorescent protein (FP) markers was proven to be problematic as significant loss of the FP-signal was routinely observed. Using splenocytes harvested from mice constitutively expressing the enhanced yellow fluorescent proteins (YFP) as a model, we modified the widely used intracellular staining protocol and successfully achieved simultaneous detection of both the nuclar proteins and YFP in T-regulatory cells. The improved protocol can be used to perform antibody-based intracellular characterization of FP-la... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531198 Thu, 01 Dec 2011 05:00:00 +0100 5531198 http://www.medworm.com/index.php?rid=5531197&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22150334%26dopt%3DAbstract Authors: Wang H, Fang J, Liang C, He M, Li Q, Chu C Abstract SiteFinding-PCR is a method for isolating flanking sequence tags (FSTs) of T-DNA insertion lines, but the efficiency needs to be improved. Here we report a computation-assisted design for the random primers used in SiteFinding- PCR. A short sequence, GCATG, was screened from the rice genome and used as the 3' end of the random primer. When applying the optimized primer for isolating FSTs from 168 transgenic rice lines, we obtained 107 specific products, including 64 FSTs. The efficiency of obtaining FSTs using the modified version of SiteFinding-PCR increased by 73.0% compared with the method previously reported (P < 0.01, µ test). We also provide computational results for several other plant species such as maize, so... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5531197 Thu, 01 Dec 2011 05:00:00 +0100 5531197 http://www.medworm.com/index.php?rid=5417892&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054538%26dopt%3DAbstract Authors: Blow NS PMID: 22054538 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417892 Tue, 01 Nov 2011 04:00:00 +0100 5417892 http://www.medworm.com/index.php?rid=5417891&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054539%26dopt%3DAbstract Authors: Dorman N PMID: 22054539 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417891 Tue, 01 Nov 2011 04:00:00 +0100 5417891 http://www.medworm.com/index.php?rid=5417890&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054540%26dopt%3DAbstract Authors: Ahern K PMID: 22054540 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417890 Tue, 01 Nov 2011 04:00:00 +0100 5417890 http://www.medworm.com/index.php?rid=5417889&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054541%26dopt%3DAbstract Authors: Perkel JM PMID: 22054541 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417889 Tue, 01 Nov 2011 04:00:00 +0100 5417889 http://www.medworm.com/index.php?rid=5417888&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054542%26dopt%3DAbstract Authors: Nybo K PMID: 22054542 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417888 Tue, 01 Nov 2011 04:00:00 +0100 5417888 http://www.medworm.com/index.php?rid=5417887&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054543%26dopt%3DAbstract Authors: Nybo K PMID: 22054543 [PubMed - as supplied by publisher] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417887 Tue, 01 Nov 2011 04:00:00 +0100 5417887 http://www.medworm.com/index.php?rid=5417886&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054544%26dopt%3DAbstract Authors: Stepanenko OV, Stepanenko OV, Shcherbakova DM, Kuznetsova IM, Turoverov KK, Verkhusha VV Abstract The diverse biochemical and photophysical properties of fluorescent proteins (FPs) have enabled the generation of a growing palette of colors, providing unique opportunities for their use in a variety of modern biology applications. Modulation of these FP characteristics is achieved through diversity in both the structure of the chromophore as well as the contacts between the chromophore and the surrounding protein barrel. Here we review our current knowledge of blue, green, and red chromophore formation in permanently emitting FPs, photoactivatable FPs, and fluorescent timers. Progress in understanding the interplay between FP structure and function has allowed the engineerin... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417886 Tue, 01 Nov 2011 04:00:00 +0100 5417886 http://www.medworm.com/index.php?rid=5417885&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054545%26dopt%3DAbstract Authors: Moniuszko G, Laska-Oberndorff A, Cristescu SM, Harren FJ, Sirko A PMID: 22054545 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417885 Tue, 01 Nov 2011 04:00:00 +0100 5417885 http://www.medworm.com/index.php?rid=5417884&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054546%26dopt%3DAbstract Authors: Gangalum RK, Jing Z, Nagaoka Y, Jiang M, Bhat SP, Bhat SP Abstract An unresolved bottleneck in bacterial artificial chromosome (BAC) transgenesis is low efficiency generation of founder mice because of suboptimal quality of the manipulated BAC DNA. Using mini-gel electrophoresis and electro-elution that circumvents CsCl(2) centrifugation, column chromatography, and resin purifications, we have used RECOCHIP, a commercially available dialysis cassette for the purification of BAC DNA that generates transgenic founders with up to 80% efficiency. PMID: 22054546 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417884 Tue, 01 Nov 2011 04:00:00 +0100 5417884 http://www.medworm.com/index.php?rid=5417883&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054547%26dopt%3DAbstract Authors: Chang YF, Lee-Chang JS, Panneerdoss S, Maclean JA, Rao MK Abstract A thorough understanding of the events during mammalian spermatogenesis requires studying specific molecular signatures of individual testicular cell populations as well as their interaction in co-cultures. However, most purification techniques to isolate specific testicular cell populations are timeconsuming, require large numbers of animals, and/or are only able to isolate a few cell types. Here we describe a cost-effective and timesaving approach that uses a single protocol to enrich multiple testicular cell populations (Sertoli, Leydig, and several spermatogenic cell populations) from as few as one mouse. Our protocol combines rigorous enzymatic digestion of seminiferous tubules with counter-current cen... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417883 Tue, 01 Nov 2011 04:00:00 +0100 5417883 http://www.medworm.com/index.php?rid=5417882&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22054548%26dopt%3DAbstract Authors: Yati A, Yati A, Dey S Abstract While several software programs exist to count bacterial colonies on a Petri plate, no suitable solution is available for quick and reliable enumeration of small, live insects. We have written a program called FlyCounter that can obtain counts from images, even if insects are highly clumped in space. We also describe a simple and inexpensive system for anesthetizing and capturing high-quality images of the small insects. Taken together, our process is fast, fully automatic, and has a low percentage of error (~1%-4% on average). Although we have tested our software on fruit flies, it should be simple to extend to other organisms of similar size. PMID: 22054548 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5417882 Tue, 01 Nov 2011 04:00:00 +0100 5417882 http://www.medworm.com/index.php?rid=5311685&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988683%26dopt%3DAbstract Authors: Blow NS PMID: 21988683 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311685 Sat, 01 Oct 2011 04:00:00 +0100 5311685 http://www.medworm.com/index.php?rid=5311684&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988684%26dopt%3DAbstract Authors: Blow N PMID: 21988684 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311684 Sat, 01 Oct 2011 04:00:00 +0100 5311684 http://www.medworm.com/index.php?rid=5311683&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988685%26dopt%3DAbstract Authors: Dorman N PMID: 21988685 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311683 Sat, 01 Oct 2011 04:00:00 +0100 5311683 http://www.medworm.com/index.php?rid=5311682&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988686%26dopt%3DAbstract Authors: Honig B PMID: 21988686 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311682 Sat, 01 Oct 2011 04:00:00 +0100 5311682 http://www.medworm.com/index.php?rid=5311681&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988687%26dopt%3DAbstract Authors: Ahern K PMID: 21988687 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311681 Sat, 01 Oct 2011 04:00:00 +0100 5311681 http://www.medworm.com/index.php?rid=5311680&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988688%26dopt%3DAbstract Authors: Perkel JM PMID: 21988688 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311680 Sat, 01 Oct 2011 04:00:00 +0100 5311680 http://www.medworm.com/index.php?rid=5311679&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988689%26dopt%3DAbstract Authors: Nybo K PMID: 21988689 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311679 Sat, 01 Oct 2011 04:00:00 +0100 5311679 http://www.medworm.com/index.php?rid=5311678&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988690%26dopt%3DAbstract Authors: Boehler RM, Graham JG, Shea LD Abstract Tissue engineering scaffolds have emerged as a powerful tool within regenerative medicine. These materials are being designed to create environments that promote regeneration through a combination of: (i) scaffold architecture, (ii) the use of scaffolds as vehicles for transplanting progenitor cells, and/or (iii) localized delivery of inductive factors or genes encoding for these inductive factors. This review describes the techniques associated with each of these components. Additionally, the immune response is increasingly recognized as a factor influencing regeneration. The immune reaction to an implant begins with an acute response to the injury and innate recognition of foreign materials, with the subsequent chronic immune respo... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311678 Sat, 01 Oct 2011 04:00:00 +0100 5311678 http://www.medworm.com/index.php?rid=5311677&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988691%26dopt%3DAbstract Authors: McConnell KI, Schweller RM, Diehl MR, Suh J Abstract High-throughput live-cell microarray technologies that facilitate combinatorial screening of genes and RNA interference (RNAi) would be invaluable in the identification of key gene expression profiles involved in complex cellular behaviors. Each spot on such a microarray can comprise a unique combination of genes or RNAi packaged into gene delivery vectors. Live target cells seeded on top of the microarrays would express the combination of genetic factors, potentially leading to phenotypic changes within cells. Here, we investigate the feasibility of using adeno-associated virus (AAV) as a gene delivery agent for such live-cell genetic microarrays. A robotic spotter was used to deposit AAV onto gamma-amino propyl silane,... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311677 Sat, 01 Oct 2011 04:00:00 +0100 5311677 http://www.medworm.com/index.php?rid=5311676&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988692%26dopt%3DAbstract Authors: Xiao Y, Jung C, Marx AD, Winkler I, Wyman C, Lebbink JH, Friedhoff P, Cristovao M Abstract The DNA mismatch repair (MMR) system recognizes and repairs errors that escaped the proofreading function of DNA polymerases. To study molecular details of the MMR mechanism, in vitro biochemical assays require specific DNA substrates carrying mismatches and strand discrimination signals. Current approaches used to generate MMR substrates are time-consuming and/or not very flexible with respect to sequence context. Here we report an approach to generate small circular DNA containing a mismatch (nanocircles). Our method is based on the nicking of PCR products resulting in single-stranded 3' overhangs, which form DNA circles after annealing and ligation. Depending on the DNA template, ... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311676 Sat, 01 Oct 2011 04:00:00 +0100 5311676 http://www.medworm.com/index.php?rid=5311675&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988693%26dopt%3DAbstract Authors: Jullien N, Herman JP Abstract We developed a new approach to prepare DNA probes for electrophoretic mobility gel shift assays that presents a number of advantages compared with the classical approach. The method relies on two complementary oligonucleotides containing the desired transcription factor binding sequence, one of them being extended at its 3' end with a universal tail that complements a third, short oligonucleotide labeled with a fluorophore or any other label. The use of this third "universal" oligonucleotide allows labeling many different probes with minimal time, effort and cost. We show that probes prepared this way are as effective and reliable as probes prepared by conventional methods. We refer to this short oligonucleotide as LUEGO for labeled universal ... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311675 Sat, 01 Oct 2011 04:00:00 +0100 5311675 http://www.medworm.com/index.php?rid=5311674&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988694%26dopt%3DAbstract Authors: Liu H, Hu J, Pan L, Wang S, He Y, Ding Y Abstract Global gene expression profiling (GGEP) plays a pivotal role in biological research. We developed an improved GGEP method called "robust ordered mRNA differential display (RoDD)" by combining mRNA differential display (DD) and complementary DNA amplified fragment length polymorphisms (cDNA-AFLP) using elaborately designed primers and a poly (dT:A) replacement technique. Redundancy was minimized by bead-based isolation and coverage was improved by using restriction enzymes that recognized 4-bp sites. This method offers the common virtues of gel-based methods along with the reliability of cDNA-AFLP. The most significant advantage of RoDD over current gel-based methods is greatly improved coverage and minimized redundancy.glob... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311674 Sat, 01 Oct 2011 04:00:00 +0100 5311674 http://www.medworm.com/index.php?rid=5311673&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988695%26dopt%3DAbstract Authors: Méndez JL, Ohana RF, Hurst R, Murphy N, Benink HN, Slater MR, Brueck C, Daniels DL, Urh M PMID: 21988695 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311673 Sat, 01 Oct 2011 04:00:00 +0100 5311673 http://www.medworm.com/index.php?rid=5311672&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21988696%26dopt%3DAbstract Authors: PMID: 21988696 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5311672 Sat, 01 Oct 2011 04:00:00 +0100 5311672 http://www.medworm.com/index.php?rid=5217512&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906031%26dopt%3DAbstract Authors: Blow N PMID: 21906031 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217512 Thu, 01 Sep 2011 04:00:00 +0100 5217512 http://www.medworm.com/index.php?rid=5217511&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906032%26dopt%3DAbstract Authors: Nathan B, Lo PC PMID: 21906032 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217511 Thu, 01 Sep 2011 04:00:00 +0100 5217511 http://www.medworm.com/index.php?rid=5217510&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906033%26dopt%3DAbstract Authors: Dorman N PMID: 21906033 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217510 Thu, 01 Sep 2011 04:00:00 +0100 5217510 http://www.medworm.com/index.php?rid=5217509&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906034%26dopt%3DAbstract Authors: Nybo K PMID: 21906034 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217509 Thu, 01 Sep 2011 04:00:00 +0100 5217509 http://www.medworm.com/index.php?rid=5217508&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906035%26dopt%3DAbstract Authors: Ahern K PMID: 21906035 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217508 Thu, 01 Sep 2011 04:00:00 +0100 5217508 http://www.medworm.com/index.php?rid=5217507&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906036%26dopt%3DAbstract Authors: Webb S PMID: 21906036 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217507 Thu, 01 Sep 2011 04:00:00 +0100 5217507 http://www.medworm.com/index.php?rid=5217506&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906037%26dopt%3DAbstract Authors: Nybo K PMID: 21906037 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217506 Thu, 01 Sep 2011 04:00:00 +0100 5217506 http://www.medworm.com/index.php?rid=5217505&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906038%26dopt%3DAbstract Authors: Vandenbroucke I, Marck HV, Verhasselt P, Thys K, Mostmans W, Dumont S, Eygen VV, Coen K, Tuefferd M, Aerssens J Abstract Ultra-deep sequencing (UDS) of amplicons is a major application for next-generation sequencing tech nologies, even more so for the 454 Genome Sequencer FLX. Especially for this application, errors that might be introduced during any of the sample processing or data analysis steps should be avoided or at least recognized, as they might lead to aberrant sequence variant calling. Since 454 pyrosequencing relies on PCR-driven target amplification, it is key to differentiate errors introduced during the amplification step from genuine minority variants. Thereto, optimal primer design is imperative because primer selection, primer dimer formation, and nonspeci... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217505 Thu, 01 Sep 2011 04:00:00 +0100 5217505 http://www.medworm.com/index.php?rid=5217504&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906039%26dopt%3DAbstract In this study, high resolu- tion melting curve (HRM) analysis is presented as a novel validation method for real-time PCR instruments. By ap- plying HRM analysis using a defined PCR amplicon and EvaGreen dye, information about the temperature accuracy and thermal homogeneity of the heating block was obtained. This pilot study shows the potential of our technique for temperature validation of real-time quantitative PCR thermal cyclers. Our data correlated well with the tempera- ture accuracy data obtained from the Mobile Temperature Acquisition System (MTAS; r2 = 0.93), which conforms to the National Institute of Standards and Technology criteria, and our method was reproducible in independent runs (r2 = 0.95). The advantages of this HRM-based method include: (i) temperature measurement und... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217504 Thu, 01 Sep 2011 04:00:00 +0100 5217504 http://www.medworm.com/index.php?rid=5217503&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906040%26dopt%3DAbstract Authors: Johansson P, Muslimovic A, Hultborn R, Fernström E, Hammarsten O Abstract Immunofluorescence quantification of γH2AX foci is a powerful approach to quantify DNA double-strand breaks induced by cancer therapy or accidental exposure to ionizing radiation. Here we report a modification to the γH2AX immunofluorescence labeling method, whereby cells are stained in-solution before being spotted and fixed onto microscope slides. Our modified method allows arraying of 16 patient samples/slide ready for foci counting in 2 h and demonstrated reliably detection of γH2AX foci in mononuclear cells prepared from patients who had undergone radiation therapy. PMID: 21906040 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217503 Thu, 01 Sep 2011 04:00:00 +0100 5217503 http://www.medworm.com/index.php?rid=5217502&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906041%26dopt%3DAbstract Authors: Henneberger R, Birch D, Bergquist P, Walter M, Anitori RP Abstract Visualization of microorganisms in soils and sediments using fluorescent dyes is a common method in microbial ecology studies, but is often ham- pered by strong nonspecific background fluorescence that can mask genu- ine cellular signals. The cyanine nucleic acid binding dyes TO-PRO-3 and TOTO-3 iodide enabled a clear detection of microbial cells in a mineral soil, while nonspecific background was greatly reduced compared with com- monly used dyes. When used as counterstains for fluorescence in situ hybrid- ization (FISH), both cyanine dyes allowed identification of microbial cells despite strong background from nonspecifically bound probes. TO-PRO-3 and TOTO-3 are easy to use and represent superior alterna... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217502 Thu, 01 Sep 2011 04:00:00 +0100 5217502 http://www.medworm.com/index.php?rid=5217501&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906042%26dopt%3DAbstract Authors: Boisselier E, Audet ML, Cantin L, Salesse C Abstract Glutathione S-transferase (GST) is widely used to prepare and purify GSTtagged fusion proteins. Although GST improves protein solubility, detergents must often be used to achieve protein solubilization from bacterial lysates. However, purification of GST by affinity chromatography cannot be achieved in the presence of even low concentrations of the detergent sodium dodecyl sulfate (SDS). Here we show that 2-methyl-2,4-pentanediol (MPD) can prevent SDS from interfering with purification of GST, thus enabling purification of proteins that require SDS to improve their solubility. PMID: 21906042 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217501 Thu, 01 Sep 2011 04:00:00 +0100 5217501 http://www.medworm.com/index.php?rid=5217500&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906043%26dopt%3DAbstract Authors: Morozumi T, Toki D, Eguchi-Ogawa T, Uenishi H Abstract Large-scale cDNA-sequencing projects require an efficient strategy for mass sequencing. Here we describe a method for sequencing pooled cDNA clones using a combination of transposon insertion and Gateway technology. Our method reduces the number of shotgun clones that are unsuitable for reconstruction of cDNA sequences, and has the advantage of reducing the total costs of the sequencing project. PMID: 21906043 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217500 Thu, 01 Sep 2011 04:00:00 +0100 5217500 http://www.medworm.com/index.php?rid=5217499&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21906044%26dopt%3DAbstract Authors: Teiling C, Pieprzyk M PMID: 21906044 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5217499 Thu, 01 Sep 2011 04:00:00 +0100 5217499 http://www.medworm.com/index.php?rid=5095671&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806546%26dopt%3DAbstract Authors: Blow NS PMID: 21806546 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095671 Sun, 31 Jul 2011 23:00:00 +0100 5095671 http://www.medworm.com/index.php?rid=5095668&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806547%26dopt%3DAbstract Authors: Lo PC, Nybo K PMID: 21806547 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095668 Sun, 31 Jul 2011 23:00:00 +0100 5095668 http://www.medworm.com/index.php?rid=5095662&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806548%26dopt%3DAbstract Authors: Dorman N PMID: 21806548 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095662 Sun, 31 Jul 2011 23:00:00 +0100 5095662 http://www.medworm.com/index.php?rid=5095657&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806549%26dopt%3DAbstract Authors: Nybo K PMID: 21806549 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095657 Sun, 31 Jul 2011 23:00:00 +0100 5095657 http://www.medworm.com/index.php?rid=5095653&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806550%26dopt%3DAbstract Authors: Ahern K PMID: 21806550 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095653 Sun, 31 Jul 2011 23:00:00 +0100 5095653 http://www.medworm.com/index.php?rid=5095649&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806551%26dopt%3DAbstract Authors: Perkel JM PMID: 21806551 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095649 Sun, 31 Jul 2011 23:00:00 +0100 5095649 http://www.medworm.com/index.php?rid=5095643&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806552%26dopt%3DAbstract Authors: Nybo K PMID: 21806552 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095643 Sun, 31 Jul 2011 23:00:00 +0100 5095643 http://www.medworm.com/index.php?rid=5095630&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806553%26dopt%3DAbstract Authors: Watson DS, Jambunathan K, Askew DS, Kodukula K, Galande AK Proteases are candidate biomarkers and therapeutic targets for many diseases. Sensitive and robust techniques are needed to quantify proteolytic activities within the complex biological milieu. We hypothesized that a combinatorial protease substrate library could be used effectively to identify similarities and differences between serum and bronchoalveolar lavage fluid (BALF), two body fluids that are clinically important for developing targeted therapies and diagnostics. We used a concise library of fluorogenic probes to map the protease substrate specificities of serum and BALF from guinea pigs. Differences in the proteolytic fingerprints of the two fluids were striking: serum proteases cleaved substrates containing ... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095630 Sun, 31 Jul 2011 23:00:00 +0100 5095630 http://www.medworm.com/index.php?rid=5095601&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806554%26dopt%3DAbstract Authors: Leippe DM, Nguyen D, Zhou M, Good T, Kirkland TA, Scurria M, Bernad L, Ugo T, Vidugiriene J, Cali JJ, Klaubert DH, O'Brien MA A bioluminescent general protease assay was developed using a combination of five luminogenic peptide substrates. The peptide-conjugated luciferin substrates were combined with luciferase to form a homogeneous, coupled-enzyme assay. This single-reagent format minimized backgrounds, gave stable signals, and reached peak sensitivity within 30 min. The bioluminescent assay was used to detect multiple proteases representing serine, cysteine, and metalloproteinase classes. The range of proteases detected was broader and the sensitivity greater, when compared with a standard fluorescent assay based on cleavage of the whole protein substrate casein. Fifteen of... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095601 Sun, 31 Jul 2011 23:00:00 +0100 5095601 http://www.medworm.com/index.php?rid=5095588&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806555%26dopt%3DAbstract Authors: Trifilieff P, Rives ML, Urizar E, Piskorowski RA, Vishwasrao HD, Castrillon J, Schmauss C, Slättman M, Gullberg M, Javitch JA The existence of G protein-coupled receptor (GPCR) dimers and/or oligomers has been demonstrated in heterologous systems using a variety of biochemical and biophysical assays. While these interactions are the subject of intense research because of their potential role in modulating signaling and altering pharmacology, evidence for the existence of receptor interactions in vivo is still elusive because of a lack of appropriate methods to detect them. Here, we adapted and optimized a proximity ligation assay (PLA) for the detection in brain slices of molecular proximity of two antigens located on either the same or two different GPCRs. Using this approac... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095588 Sun, 31 Jul 2011 23:00:00 +0100 5095588 http://www.medworm.com/index.php?rid=5095533&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806556%26dopt%3DAbstract Authors: Mahon MJ The simian virus 40 large T antigen (SVLT) induces replication of plasmids bearing the SV40 origin of replication (SV40 ori) within mammalian cells. The internal ribosomal entry site (IRES) is an element that allows for the cotranslation of proteins from one polycistronic mRNA. Through the combination of these elements, IRES-dependent coexpression of a protein of interest and the SVLT, either constitutive or regulated, on plasmids bearing the SV40 ori generates a positive feedback loop, resulting in enhanced expression. A vector linking red fluorescent protein (RFP) to the IRES-SVLT element enhances fluorescence ~10-fold over that demonstrated from a vector lacking this element. In transfection-resistant CV-1 cells, the RFP-IRES-SVLT vector substantially increases the... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095533 Sun, 31 Jul 2011 23:00:00 +0100 5095533 http://www.medworm.com/index.php?rid=5095499&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21806557%26dopt%3DAbstract Authors: Fordyce SL, Avila-Arcos MC, Rockenbauer E, Børsting C, Frank-Hansen R, Petersen FT, Willerslev E, Hansen AJ, Morling N, Gilbert MT The analysis and profiling of short tandem repeat (STR) loci is routinely used in forensic genetics. Current methods to investigate STR loci, including PCR-based standard fragment analyses and capillary electrophoresis, only provide amplicon lengths that are used to estimate the number of STR repeat units. These methods do not allow for the full resolution of STR base composition that sequencing approaches could provide. Here we present an STR profiling method based on the use of the Roche Genome Sequencer (GS) FLX to simultaneously sequence multiple core STR loci. Using this method in combination with a bioinformatic tool designed specifically to... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095499 Sun, 31 Jul 2011 23:00:00 +0100 5095499 http://www.medworm.com/index.php?rid=5095733&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781043%26dopt%3DAbstract Authors: Blow NS PMID: 21781043 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095733 Thu, 30 Jun 2011 23:00:00 +0100 5095733 http://www.medworm.com/index.php?rid=5095723&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781044%26dopt%3DAbstract Authors: Lo PC, Nybo K PMID: 21781044 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095723 Thu, 30 Jun 2011 23:00:00 +0100 5095723 http://www.medworm.com/index.php?rid=5095718&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781045%26dopt%3DAbstract Authors: Dorman N PMID: 21781045 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095718 Thu, 30 Jun 2011 23:00:00 +0100 5095718 http://www.medworm.com/index.php?rid=5095710&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781046%26dopt%3DAbstract Authors: Nybo K PMID: 21781046 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095710 Thu, 30 Jun 2011 23:00:00 +0100 5095710 http://www.medworm.com/index.php?rid=5095708&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781047%26dopt%3DAbstract Authors: Ahern K PMID: 21781047 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095708 Thu, 30 Jun 2011 23:00:00 +0100 5095708 http://www.medworm.com/index.php?rid=5095704&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781048%26dopt%3DAbstract Authors: Perkel JM PMID: 21781048 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095704 Thu, 30 Jun 2011 23:00:00 +0100 5095704 http://www.medworm.com/index.php?rid=5095698&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781049%26dopt%3DAbstract Authors: Nybo K PMID: 21781049 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095698 Thu, 30 Jun 2011 23:00:00 +0100 5095698 http://www.medworm.com/index.php?rid=5095696&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781050%26dopt%3DAbstract Authors: Yang Y, Lin J, Meschewski R, Watson E, Valentine MT We have designed and built a magnetic tweezers device that enables the application of calibrated stresses to soft materials while simultaneously measuring their microscale deformation using confocal microscopy. Unlike previous magnetic tweezers designs, our device is entirely portable, allowing easy use on microscopes in core imaging facilities or in collaborators' laboratories. The imaging capabilities of the microscope are unimpaired, enabling the 3-D structures of fluorescently labeled materials to be precisely determined under applied load. With this device, we can apply a large range of forces (~1-1200 pN) over micron-scale contact areas to beads that are either embedded within 3-D matrices or attached to the surface of ... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095696 Thu, 30 Jun 2011 23:00:00 +0100 5095696 http://www.medworm.com/index.php?rid=5095688&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781051%26dopt%3DAbstract Authors: Ranall MV, Gabrielli BG, Gonda TJ Neutral lipid droplets (LDs) are dynamic lipid storage organelles found in all eukaryotic cells from yeast to mammals and higher plants. LDs are important to many physiological processes that include basic cellular maintenance, metabolism, and diverse medical pathologies. LD accumulation has been studied extensively by a range of methods, but particularly by microscopy with several fluorescent dyes extensively used for qualitative and quantitative imaging. Here, we compared established LD stains Nile Red and BODIPY 493/503 to the 4', 6-diamidino-2-phenylindole (DAPI)-range dye 1,6-diphenyl-1,3,5-hexatriene (DPH; excitation/emission λmax=350 nm/420 nm) using high-content image analysis. HeLa cells treated with oleic acid or vehicle were used t... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095688 Thu, 30 Jun 2011 23:00:00 +0100 5095688 http://www.medworm.com/index.php?rid=5095686&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781052%26dopt%3DAbstract Authors: Anderson EK, Martin DS Fluorescent imaging of cytoskeletal structures permits studies of both organization within the cell and dynamic reorganization of the cytoskeleton itself. Traditional fluorescent labels of microtubules, part of the cytoskeleton, have been used to study microtubule localization, structure, and dynamics, both in vivo and in vitro. However, shortcomings of existing labels make imaging of microtubules with high precision light microscopy difficult. In this paper, we report a new fluorescent labeling technique for microtubules, which involves a GTP analog modified with a bright, organic fluorophore (TAMRA, Cy3, or Cy5). This fluorescent GTP binds to a specific site, the exchangeable site, on tubulin in solution with a dissociation constant of 1.0±0.4 µM. Fu... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095686 Thu, 30 Jun 2011 23:00:00 +0100 5095686 http://www.medworm.com/index.php?rid=5095684&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781053%26dopt%3DAbstract Authors: Puah WC, Cheok LP, Biro M, Ng WT, Wasser M Automated microscopy enables in vivo studies in developmental biology over long periods of time. Time-lapse recordings in three or more dimensions to study the dynamics of developmental processes can produce huge data sets that extend into the terabyte range. However, depending on the available computational resources and software design, downstream processing of very large image data sets can become highly inefficient, if not impossible. To address the lack of available open source and commercial software tools to efficiently reorganize time-lapse data on a desktop computer with limited system resources, we developed TLM-Converter. The software either fragments oversized files or concatenates multiple files representing single time f... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095684 Thu, 30 Jun 2011 23:00:00 +0100 5095684 http://www.medworm.com/index.php?rid=5095681&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781054%26dopt%3DAbstract Authors: Matsumoto A, Itoh TQ Enzyme-free cloning (EFC) can rapidly produce an in-frame fusion gene with multiple fragments. To practically apply EFC, we investigated the extent and sequence of complementary staggered overhangs necessary to direct self-assembly of multiple fragments as well as a size limitation of the constructed DNA molecule. Six-base pair overhangs with 50% GC content were sufficient to direct self-assembly. A functional plasmid that exceeded 10 kb, which includes an in-frame fusion domain, was efficiently constructed from four PCR fragments in one step by our improved method. PMID: 21781054 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095681 Thu, 30 Jun 2011 23:00:00 +0100 5095681 http://www.medworm.com/index.php?rid=5095675&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781055%26dopt%3DAbstract Authors: Benko Z, Zhao RY Complementation of auxotrophic nutrient deficiencies in minimal media is widely used for selection of exogenous gene introduction to fission yeast. However, only a limited number of such selection markers are available. Antibiotic resistance markers are good alternatives, but they typically work well in complete rich medium but not in minimal defined Edinburgh minimal medium (EMM). It would be ideal if both the auxotrophic and antibiotic resistance markers can be used together for molecular genetic analysis. Here we describe the use of Zeocin in Pombe minimal glutamate (PMG) media for selection and maintenance of bleMX6 resistance with a LEU2 auxotrophic marker in fission yeast. PMID: 21781055 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095675 Thu, 30 Jun 2011 23:00:00 +0100 5095675 http://www.medworm.com/index.php?rid=5095792&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781029%26dopt%3DAbstract Authors: Blow NS PMID: 21781029 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095792 Tue, 31 May 2011 23:00:00 +0100 5095792 http://www.medworm.com/index.php?rid=5095790&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781030%26dopt%3DAbstract Authors: Lo PC, Nybo K PMID: 21781030 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095790 Tue, 31 May 2011 23:00:00 +0100 5095790 http://www.medworm.com/index.php?rid=5095788&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781031%26dopt%3DAbstract Authors: Dorman N PMID: 21781031 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095788 Tue, 31 May 2011 23:00:00 +0100 5095788 http://www.medworm.com/index.php?rid=5095786&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781032%26dopt%3DAbstract Authors: Nybo K PMID: 21781032 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095786 Tue, 31 May 2011 23:00:00 +0100 5095786 http://www.medworm.com/index.php?rid=5095784&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781033%26dopt%3DAbstract Authors: Ahern K PMID: 21781033 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095784 Tue, 31 May 2011 23:00:00 +0100 5095784 http://www.medworm.com/index.php?rid=5095782&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781034%26dopt%3DAbstract Authors: Perkel JM PMID: 21781034 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095782 Tue, 31 May 2011 23:00:00 +0100 5095782 http://www.medworm.com/index.php?rid=5095777&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781035%26dopt%3DAbstract Authors: Nybo K PMID: 21781035 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095777 Tue, 31 May 2011 23:00:00 +0100 5095777 http://www.medworm.com/index.php?rid=5095763&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781036%26dopt%3DAbstract Authors: PMID: 21781036 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095763 Tue, 31 May 2011 23:00:00 +0100 5095763 http://www.medworm.com/index.php?rid=5095761&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781037%26dopt%3DAbstract Authors: Tran TN, Aboudharam G, Raoult D, Drancourt M Identifying the causes of past epidemics depends on the specific detection of pathogens in buried individuals; this field of research is known as paleomicrobiology, an emerging field that has benefited from technological advances in microbiology. For almost 15 years, the detection, identification, and characterization of microbes in ancient environmental and human specimens emerged on the basis of ancient DNA (aDNA) analyses. aDNA limitations due to potential contamination by modern DNA and altered aDNA led to the development of alternative methods for the detection and characterization of nonnucleotidic biomolecules, including mycolic acids (of ancient mycobacteria) and proteins. Accordingly, immunohistochemistry, immunochromatogra... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095761 Tue, 31 May 2011 23:00:00 +0100 5095761 http://www.medworm.com/index.php?rid=5095759&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781038%26dopt%3DAbstract In this study, we immobilized mouse stem cells and non-stem cells (control) on a glass surface as a microarray and reacted the cell microarray with unlabeled SSEA1 antibodies. By monitoring the reaction with an OI-RD microscope in real time, we confirmed that the SSEA1 antibodies bind only to the surface of the stem cells and not to the surface of non-stem cells. From the binding curves, we determined the equilibrium dissociation constant (Kd) of the antibody with the SSEA1 markers on the stem cell surface. Thus, the OI-RD microscope can be used to detect binding affinities between cell surface markers and unlabeled antibodies bound to the cells; this information could be useful for determination of stem cell stages. PMID: 21781038 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095759 Tue, 31 May 2011 23:00:00 +0100 5095759 http://www.medworm.com/index.php?rid=5095757&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781039%26dopt%3DAbstract Authors: Pelts M, Pandya SM, Oh CJ, Model MA Conventional light microscopy techniques are poorly suited for imaging the vertical cell dimension. This can be accomplished using transmission-through-dye (TTD) imaging, in which cell thickness is directly converted into image intensity in the presence of extracellular dye with strong absorption. We have previously described applications of TTD to living cells using the dye Acid Blue 9 (AB9) to generate contrast. In this work, we investigated the possibility of extending TTD to chemically fixed cells. This would depend on preservation of cell impermeability to the dye; by using a method based on fluorescence quenching, we found that formaldehyde-fixed cells remain impermeable to AB9. Fixation enables imaging of cell surfaces in the presence... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095757 Tue, 31 May 2011 23:00:00 +0100 5095757 http://www.medworm.com/index.php?rid=5095749&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781040%26dopt%3DAbstract Authors: Zylicz-Stachula A, Zołnierkiewicz O, Jeżewska-Frąckowiak J, Skowron PM The type IIS/IIC restriction endonuclease TspGWI recognizes the sequence 5'-ACGGA-3', cleaving DNA 11/9 nucleotides downstream. Here we show that sinefungin, a cofactor analog of S-adenosyl methionine, induces a unique type of relaxation in DNA recognition specificity. In the presence of sinefungin, TspGWI recognizes and cleaves at least 12 degenerate variants of the original recognition sequence that vary by single base pair changes from the original 5-bp restriction site with only a single degeneracy per variant appearing to be allowed. In addition, sinefungin was found to have a stimulatory effect on cleavage at these nondegenerate TspGWI recognition sites, irrespective of their number or the DNA topo... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095749 Tue, 31 May 2011 23:00:00 +0100 5095749 http://www.medworm.com/index.php?rid=5095742&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781041%26dopt%3DAbstract Authors: Ortega LM, Hengartner CJ, Vega LR Because of their low abundance and short length, telomeric single-stranded extensions have not traditionally been assessed by Southern blot analysis. Instead, most methods have relied on hybridizing radioactively labeled oligonucleotide probes to electrophoresed DNA within agarose gels. Here we describe a rapid and nonradioactive Southern blot-derived method to transfer and detect telomeric single-stranded G-rich overhangs (G-tails) under nondenaturing (native) conditions, using Saccharomyces cerevisiae DNA. Restriction enzyme-digested chromosomal DNA is separated by agarose gel electrophoresis, transferred onto a charged membrane by electroblotting under nondenaturing conditions, and probed with a digoxigenin (DIG)-labeled oligonucleotide. Co... BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095742 Tue, 31 May 2011 23:00:00 +0100 5095742 http://www.medworm.com/index.php?rid=5095738&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21781042%26dopt%3DAbstract We describe YGFP, a slow bleaching green fluorescent protein (GFP) with unique spectral properties. YGFP is derived from an Escherichia coli codon-optimized synthetic gfp mutant 2 derivative. In addition to the GFP-mut 2 changes, it also carries S202F and T203I substitutions. YGFP can be used as a substitute for yellow fluorescent protein (YFP) in experiments in which two or more fluorescent proteins are fused to different cellular protein components, expanding the ability to study multiple labeled proteins in a cell at once. PMID: 21781042 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=5095738 Tue, 31 May 2011 23:00:00 +0100 5095738 http://www.medworm.com/index.php?rid=4855430&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21548905%26dopt%3DAbstract Authors: Perkel JM PMID: 21548905 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4855430 Thu, 31 Mar 2011 23:00:00 +0100 4855430 http://www.medworm.com/index.php?rid=4855426&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21548906%26dopt%3DAbstract We describe a versatile and rapid approach to quantitatively assay macrophage burden in intact tissue samples. Perfluorocarbon (PFC) emulsion is injected intravenously, and the emulsion droplets are effectively taken up by monocytes and macrophages. These 'in situ' labeled cells participate in inflammatory events in vivo resulting in PFC accumulation at inflammatory loci. Necropsied tissues or intact organs are subjected to conventional fluorine-19 ((19)F) NMR spectroscopy to quantify the total fluorine content per sample, proportional to the macrophage burden. We applied these methods to a rat model of experimental allergic encephalomyelitis (EAE) exhibiting extensive inflammation and demyelination in the central nervous system (CNS), particularly in the spinal cord. In a cohort of EAE ra... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4855426 Thu, 31 Mar 2011 23:00:00 +0100 4855426 http://www.medworm.com/index.php?rid=4855412&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21548907%26dopt%3DAbstract In this study, a site-specific recombination assay in Escherichia coli was used to examine the activity of mutant VCre (H314L and Y349F) and mutant SCre (H317L and Y352F), in which both mutated residues lie within the active center of Cre recombination. The site-specific recombination activity of both mutants was significantly decreased. Recombinase-mediated cassette exchange (RMCE) using VloxP and the Vlox2272 mutant site was performed in E. coli by introducing a cassette bearing VloxP and Vlox2272 into a recipient plasmid bearing the same sites. RMCE using SloxP and Slox2272 was also performed by SCre recombinase. Moreover, BAC engineering via Red recombination and VCre/VloxP were demonstrated. First, the DNA cassette for modification was introduced into a BAC clone via Red recombination... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4855412 Thu, 31 Mar 2011 23:00:00 +0100 4855412 http://www.medworm.com/index.php?rid=4855410&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21548908%26dopt%3DAbstract Authors: Vela D, Guerreiro MP, Fontdevila A An adapted amplified fragment length polymorphism (AFLP) protocol is presented for detection of hybrid instability in the genome of interspecific hybrids between Drosophila buzzatii and D. koepferae species. Analyses of 15 AFLP instability markers (new bands detected in hybrids) show that up to 81% are the result of transposable element (TE) activity. Twenty TEs associated with AFLP instability markers have been detected by this method in backcross hybrids and segmental hybrids, demonstrating its validity in detecting transposition events occurring during the hybridization process. New insertions of Helena TE have been observed in the hybrid genome after hybridization of the TGTCG22 instability marker by FISH. The AFLP marker technique proved... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4855410 Thu, 31 Mar 2011 23:00:00 +0100 4855410 http://www.medworm.com/index.php?rid=4855396&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21548909%26dopt%3DAbstract Authors: Sigot V, Pellegrino JM, Guibert EE, Rodriguez JV An inexpensive modular perfused chamber (MPC) designed for low- and normal-temperature live-cell imaging is presented. The device consists of four lathed pieces of stainless steel assembled as a cylindrical open chamber that can hold either round or square glass coverslips. The chamber is connected to a thermal-bath operating with recirculation. For image acquisition at 4°C, cooled air is blown toward the coverslip surface to prevent condensation. Principal advantages of this device are thermal stability in the sample environment, rapid response to changes in temperature set point, and easy sample insertion. The device enables the study of dynamic processes in cells governed by large temperature differences such as those impose... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4855396 Thu, 31 Mar 2011 23:00:00 +0100 4855396 http://www.medworm.com/index.php?rid=4855329&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21548910%26dopt%3DAbstract We report here an improved method for analyzing protein surface expression utilizing a cold-adapted trypsin. Preservation of activity of the enzyme at 0-4°C permits modification of the protease method of surface analysis to temperatures at which trafficking of mammalian plasmalemmal proteins is blocked. This is an important advantage over established trypsin-cleavage protocols. Moreover, the method is less time-consuming than surface biotinylation. PMID: 21548910 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4855329 Thu, 31 Mar 2011 23:00:00 +0100 4855329 http://www.medworm.com/index.php?rid=4855323&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21548911%26dopt%3DAbstract Authors: Xing M, Shen H, Zhao W, Liu Y, Du Y, Yu Z, Chen X Because of their unique spectral properties, quantum dots (QDs) have recently proved useful as fluorescent labels for biosensing probes. We developed a versatile QD label by modifying dsDNA with biotin and thiol groups at opposite ends and attaching it to quantum dots via a metal-thiol bond. These dsDNA-coated QDs fluorescently label their targets through biotin-streptavidin binding and show excellent histological results when used to detect biotin-labeled chromosome probes. The dsDNA coating also circumvented the common problems of aggregation and steric hindrance that occur with other QD probes. PMID: 21548911 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4855323 Thu, 31 Mar 2011 23:00:00 +0100 4855323 http://www.medworm.com/index.php?rid=4905436&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21626735%26dopt%3DAbstract Authors: PMID: 21626735 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4905436 Tue, 01 Mar 2011 00:00:00 +0100 4905436 http://www.medworm.com/index.php?rid=4905366&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21626736%26dopt%3DAbstract Authors: PMID: 21626736 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4905366 Tue, 01 Mar 2011 00:00:00 +0100 4905366 http://www.medworm.com/index.php?rid=4802299&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486229%26dopt%3DAbstract Authors: Blow NS PMID: 21486229 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802299 Tue, 01 Mar 2011 00:00:00 +0100 4802299 http://www.medworm.com/index.php?rid=4802298&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486230%26dopt%3DAbstract Authors: Lo PC, Nybo K PMID: 21486230 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802298 Tue, 01 Mar 2011 00:00:00 +0100 4802298 http://www.medworm.com/index.php?rid=4802297&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486231%26dopt%3DAbstract Authors: Dorman N PMID: 21486231 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802297 Tue, 01 Mar 2011 00:00:00 +0100 4802297 http://www.medworm.com/index.php?rid=4802296&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486232%26dopt%3DAbstract Authors: Nybo K PMID: 21486232 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802296 Tue, 01 Mar 2011 00:00:00 +0100 4802296 http://www.medworm.com/index.php?rid=4802295&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486233%26dopt%3DAbstract Authors: Ahern K PMID: 21486233 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802295 Tue, 01 Mar 2011 00:00:00 +0100 4802295 http://www.medworm.com/index.php?rid=4802294&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486234%26dopt%3DAbstract Authors: Perkel JM PMID: 21486234 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802294 Tue, 01 Mar 2011 00:00:00 +0100 4802294 http://www.medworm.com/index.php?rid=4802293&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486235%26dopt%3DAbstract Authors: Nybo K PMID: 21486235 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802293 Tue, 01 Mar 2011 00:00:00 +0100 4802293 http://www.medworm.com/index.php?rid=4802292&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486236%26dopt%3DAbstract Authors: Reins J, Mossner M, Richter L, Kmetsch A, Thiel E, Haase D, Hofmann WK PMID: 21486236 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802292 Tue, 01 Mar 2011 00:00:00 +0100 4802292 http://www.medworm.com/index.php?rid=4802291&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486237%26dopt%3DAbstract Authors: Liu H, McNicol J, Bayer M, Morris JA, Cardle L, Marshall DF, Schulte D, Stein N, Shi BJ, Taudien S, Waugh R, Hedley PE Second-generation sequencing now provides the potential for low-cost generation of whole-genome sequences. However, for large-genome organisms with high repetitive DNA content, genome-wide short read sequence assembly is currently impossible, with accurate ordering and localization of genes still relying heavily on integration with physical and genetic maps. To facilitate this process, we have used Agilent microarrays to simultaneously address thousands of gene sequences to individual BAC clones and contiguous sequences that form part of an emerging physical map of the large and currently unsequenced 5.3-Gb barley genome. The approach represents a cost-effecti... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802291 Tue, 01 Mar 2011 00:00:00 +0100 4802291 http://www.medworm.com/index.php?rid=4802290&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486238%26dopt%3DAbstract In this study, we tested the NuGEN Ovation RNA-Seq System for library preparation followed by next-generation sequencing on an Illumina GAIIx. The cDNA product of the NuGEN kit may have significant amounts of ssDNA with hairpin structures that are generated during the amplification process. These structures interfere with efficient downstream end repair, A-tailing, and adapter ligation, all standard steps in post-amplification sequencing library construction. We were able to increase the efficiency of sequencing library yields 4- to 6-fold or greater by treatment of NuGENamplified cDNA product with the single-strand endonuclease S1. These results suggest that this treatment effectively cleaves hairpin structures generated during amplification that are resistant to the standard enzyme cockt... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802290 Tue, 01 Mar 2011 00:00:00 +0100 4802290 http://www.medworm.com/index.php?rid=4802289&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486239%26dopt%3DAbstract Authors: Holloway TP, Rowley SM, Delatycki MB, Sarsero JP Friedreich ataxia is a neurodegenerative disorder caused by the expansion of a GAA trinucleotide repeat sequence within the first intron of the FXN gene. Interruptions in the GAA repeat may serve to alleviate the inhibitory effects of the GAA expansion on FXN gene expression and to decrease pathogenicity. We have developed a simple and rapid PCR- and restriction enzyme-based assay to asess the purity of GAA repeat sequences. PMID: 21486239 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802289 Tue, 01 Mar 2011 00:00:00 +0100 4802289 http://www.medworm.com/index.php?rid=4802288&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486240%26dopt%3DAbstract We describe a simple and rapid magnetofection-based method suitable for the lab bench as well as for high-throughput projects. Our method yields high transfection efficiency and can be used for deciphering the genetic control of neural cell differentiation. PMID: 21486240 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802288 Tue, 01 Mar 2011 00:00:00 +0100 4802288 http://www.medworm.com/index.php?rid=4802287&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486241%26dopt%3DAbstract Authors: PMID: 21486241 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802287 Tue, 01 Mar 2011 00:00:00 +0100 4802287 http://www.medworm.com/index.php?rid=4802286&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486242%26dopt%3DAbstract Authors: Blow N PMID: 21486242 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802286 Tue, 01 Feb 2011 00:00:00 +0100 4802286 http://www.medworm.com/index.php?rid=4802285&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486243%26dopt%3DAbstract Authors: Lo PC, Nybo K PMID: 21486243 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802285 Tue, 01 Feb 2011 00:00:00 +0100 4802285 http://www.medworm.com/index.php?rid=4802284&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486244%26dopt%3DAbstract Authors: Dorman N PMID: 21486244 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802284 Tue, 01 Feb 2011 00:00:00 +0100 4802284 http://www.medworm.com/index.php?rid=4802283&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486245%26dopt%3DAbstract Authors: Nybo K PMID: 21486245 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802283 Tue, 01 Feb 2011 00:00:00 +0100 4802283 http://www.medworm.com/index.php?rid=4802282&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486246%26dopt%3DAbstract Authors: Ahern K PMID: 21486246 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802282 Tue, 01 Feb 2011 00:00:00 +0100 4802282 http://www.medworm.com/index.php?rid=4802281&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486247%26dopt%3DAbstract Authors: Nybo K PMID: 21486247 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802281 Tue, 01 Feb 2011 00:00:00 +0100 4802281 http://www.medworm.com/index.php?rid=4802280&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486248%26dopt%3DAbstract Authors: PMID: 21486248 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802280 Tue, 01 Feb 2011 00:00:00 +0100 4802280 http://www.medworm.com/index.php?rid=4802279&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486249%26dopt%3DAbstract Authors: Perkel J PMID: 21486249 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802279 Tue, 01 Feb 2011 00:00:00 +0100 4802279 http://www.medworm.com/index.php?rid=4802278&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486250%26dopt%3DAbstract Authors: Smith HE Insertion mutagenesis via mobile genetic element is a common technique for the analysis of gene function in model organisms. Next-generation sequencing offers an attractive approach for localizing the site of insertion, but alignment-based mapping of mobile genetic elements is challenging. A computational method for identifying insertion sites is reported herein. The technique was validated by mapping transposons in both bacterial and nematode species. The approach should be extensible to other systems that employ mobile genetic elements to generate mutations. PMID: 21486250 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802278 Tue, 01 Feb 2011 00:00:00 +0100 4802278 http://www.medworm.com/index.php?rid=4802277&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486251%26dopt%3DAbstract Authors: Perry SW, Norman JP, Barbieri J, Brown EB, Gelbard HA Fluorescent probes for monitoring mitochondrial membrane potential are frequently used for assessing mitochondrial function, particularly in the context of cell fate determination in biological and biomedical research. However, valid interpretation of results obtained with such probes requires careful consideration of numerous controls, as well as possible effects of non-protonic charges on dye behavior. In this context, we provide an overview of some of the important technical considerations, controls, and parallel complementary assays that can be employed to help ensure appropriate interpretation of results, thus providing a practical usage guide for monitoring mitochondrial membrane potentials with cationic probes. In to... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802277 Tue, 01 Feb 2011 00:00:00 +0100 4802277 http://www.medworm.com/index.php?rid=4802276&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486252%26dopt%3DAbstract This study tested the capacity for a novel acoustic microstreaming method ("micromixing"), which stirs fluid at microliter scales, to improve cDNA yields from reverse transcription (RT) reactions comprising single-cell quantities of RNA. Micromixing significantly decreased the number of qPCR cycles to detect cDNA representing mRNA for hypoxanthine phosphoribosyl-transferase (Hprt) and nuclear receptor-related 1 (Nurr1) by ~9 and ~15 cycles, respectively. The improvement was equivalent to performing RT with 10- to 100-fold more cDNA in the absence of micromixing. Micromixing enabled reliable detection of the otherwise undetectable, low-abundance transcript, Nurr1. It was most effective when RNA concentrations were low (0.1-1 pg/µL, a "single-cell equivalent") but had lesser effects at high... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4802276 Tue, 01 Feb 2011 00:00:00 +0100 4802276 http://www.medworm.com/index.php?rid=4748476&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486253%26dopt%3DAbstract Authors: Aulanier AL, Doiron AL, Shepherd RD, Rinker KD, Frayne R, Andersen LB To determine the initial feasibility of using magnetic resonance (MR) imaging to detect early atherosclerosis, we investigated inflammatory cells labeled with a positive contrast agent in an endothelial cell-based testing system. The human monocytic cell line THP-1 was labeled by overnight incubation with a gadolinium colloid (Gado CELLTrack) prior to determination of the in vitro release profile from T1-weighted MR images. Next, MR signals arising from both a synthetic model of THP-1/human umbilical vein endothelial cell (HUVEC) accumulation and the dynamic adhesion of THP-1 cells to activated HUVECs under flow were obtained. THP-1 cells were found to be successfully-but not optimally-labeled with gadoliniu... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4748476 Tue, 01 Feb 2011 00:00:00 +0100 4748476 http://www.medworm.com/index.php?rid=4748468&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21486254%26dopt%3DAbstract Authors: Yehezkel TB, Nagar S, Mackrants D, Marx Z, Linshiz G, Shabi U, Shapiro E Bacterial cloning was first introduced over a century ago and has since become one of the most useful procedures in biological research, perhaps paralleled in its ubiquity only by PCR and DNA sequencing. However, unlike PCR and sequencing, cloning has generally remained a manual, labor-intensive, low-throughput procedure. Here we address this issue by developing an automated, computer-aided bacterial cloning method using liquid medium that is based on the principles of (i) limiting dilution of bacteria, (ii) inference of colony forming units (CFUs) based on optical density (OD) readings, and (iii) verification of monoclonality using a mixture of differently colored fluorescently labeled bacteria for trans... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4748468 Tue, 01 Feb 2011 00:00:00 +0100 4748468 http://www.medworm.com/index.php?rid=4466366&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21284241%26dopt%3DAbstract Authors: Blow NS PMID: 21284241 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4466366 Sat, 01 Jan 2011 00:00:00 +0100 4466366 http://www.medworm.com/index.php?rid=4466365&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21284242%26dopt%3DAbstract Authors: Cowan C PMID: 21284242 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4466365 Sat, 01 Jan 2011 00:00:00 +0100 4466365 http://www.medworm.com/index.php?rid=4466364&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21284243%26dopt%3DAbstract Authors: Blow NS PMID: 21284243 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4466364 Sat, 01 Jan 2011 00:00:00 +0100 4466364 http://www.medworm.com/index.php?rid=4399766&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21231919%26dopt%3DAbstract Authors: Beisvåg V, Kauffmann A, Malone J, Foy C, Salit M, Schimmel H, Bongcam-Rudloff E, Landegren U, Parkinson H, Huber W, Brazma A, Sandvik A, Kuiper M While minimum information about a microarray experiment (MIAME) standards have helped to increase the value of the microarray data deposited into public databases like ArrayExpress and Gene Expression Omnibus (GEO), limited means have been available to assess the quality of this data or to identify the procedures used to normalize and transform raw data. The EMERALD FP6 Coordination Action was designed to deliver approaches to assess and enhance the overall quality of microarray data and to disseminate these approaches to the microarray community through an extensive series of workshops, tutorials, and symposia. Tools were developed... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4399766 Sat, 01 Jan 2011 00:00:00 +0100 4399766 http://www.medworm.com/index.php?rid=4399765&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21231920%26dopt%3DAbstract Authors: Kodoyianni V Surface plasmon resonance (SPR) is a label-free detection method by which molecular interactions may be analyzed on a surface. Binding data are collected in real time, allowing the determination of interaction kinetics. SPR imaging (SPRi), the focus of this review, improves upon the efficiency of SPR by facilitating analysis of multiple interactions simultaneously. Here we summarize the principles of SPRi, provide examples of how SPRi arrays can be fabricated, and illustrate the utility of SPRi through example applications from the fields of proteomics, genomics and bioengineering. Address correspondence to Voula Kodoyianni, GWC Technologies Inc., 505 South Rosa Road, Madison, WI, 53719, USA. e-mail: PMID: 21231920 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4399765 Sat, 01 Jan 2011 00:00:00 +0100 4399765 http://www.medworm.com/index.php?rid=4399764&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21231921%26dopt%3DAbstract Authors: Forrester M, Eyler C, Rich J A wide range of mammalian signaling and stress pathways are mediated by nitric oxide (NO), which is synthesized in vivo by the nitric oxide synthase (NOS) family of enzymes. Experimental manipulations of NO are frequently achieved by either inhibition or activation of endogenous NOS or via providing exogenous NO sources. On the contrary, many microbes consume NO via flavohemoglobin (FlavoHb), a highly efficient NO-dioxygenase that protects from nitrosative stress. Here we report a novel resource for studying NO in mammalian cells by heterologously expressing Escherichia coli FlavoHb within a lentiviral delivery system. This technique boosts endogenous cellular consumption of NO, thus providing a simple and efficacious approach to studying mammalian... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4399764 Sat, 01 Jan 2011 00:00:00 +0100 4399764 http://www.medworm.com/index.php?rid=4399763&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21231922%26dopt%3DAbstract Authors: Callejas S, Alvarez R, Dopazo A Quantitative PCR (qPCR) remains the method of choice for gene and microRNA (miRNA) expression studies. Many laboratories wish to automate some or all of the steps of medium-throughput qPCR experiments through the use of various types of liquid handling robots. However, it is not uncommon to find cases in which scripts provided by the robot supplier are too rigid for user-specific applications, do not include all the desired options, or are too complicated to be modified by a nonprofessional programmer. Here, we present Automatic Genomics, a program that allows users with a limited programming background to automate medium-throughput qPCR experiments by using commercially available liquid-handling robots. The user is able to optimize the plate de... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4399763 Sat, 01 Jan 2011 00:00:00 +0100 4399763 http://www.medworm.com/index.php?rid=4399762&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21231923%26dopt%3DAbstract Authors: Chou WP, Chen PH, Miao M, Kuo LS, Yeh SH, Chen PJ Herein we describe a simple platform for rapid DNA amplification using convection. Capillary convective PCR (CCPCR) heats the bottom of a capillary tube using a dry bath maintained at a fixed temperature of 95°C. The tube is then cooled by the surrounding air, creating a temperature gradient in which a sample can undergo PCR amplification by natural convection through reagent circulation. We demonstrate that altering the melting temperature of the primers relative to the lowest temperature in the tube affects amplification efficiency; adjusting the denaturation temperature of the amplicon relative to the highest temperature in the tube affects maximum amplicon size, with amplicon lengths of ≤500 bp possible. Based on these c... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4399762 Sat, 01 Jan 2011 00:00:00 +0100 4399762 http://www.medworm.com/index.php?rid=4399761&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21231924%26dopt%3DAbstract Authors: Kodedová M, Sigler K, Lemire B, Gášková D New antifungal agents are needed to treat life-threatening fungal infections, particularly with the development of resistance. Surface-active antifungals have the advantages of minimizing host toxicity and the emergence of drug resistance. We have developed a time-dependent drug exposure assay that allows us to rapidly investigate the mechanism of surface-active antifungal drug action. The assay uses a multidrug pump-deficient strain of Saccharomyces cerevisiae and the potentiometric dye 3,3'-dipropylthiacarbocyanine iodide [diS-C3(3)] and can assess whether cells are depolarized, hyperpolarized, or permeabilized by drug exposure. In this work, we investigated the mechanisms of action of five surface-active compounds: SDS, nystatin... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4399761 Sat, 01 Jan 2011 00:00:00 +0100 4399761 http://www.medworm.com/index.php?rid=4273923&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21143208%26dopt%3DAbstract Authors: Perkel J PMID: 21143208 [PubMed - in process] (Source: BioTechniques) BioTechniques journals http://www.medworm.com/rss/comments.php?id=4273923 Wed, 01 Dec 2010 00:00:00 +0100 4273923 http://www.medworm.com/index.php?rid=4273922&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21143209%26dopt%3DAbstract Authors: Huang C, Jacobson K Detection of protein-protein interactions in cells is crucial for understanding the biological functions of proteins, including their roles in signal transduction. However, current methods require specific antibodies both for immunoprecipitation and detection, making them expensive and sometimes unreliable. Here we describe protocols for protein-protein interaction assays that use nonimmune IgG-conjugated Sepharose to precipitate the IgG binding domain (ZZ) fused to the bait protein; the interaction partner is fused to Avitag and biotinylated by BirA so that it can be detected by a one-step blot with Dylight 680 streptavidin to detect the Avitag fusion protein. Since this method does not require specific antibodies and is inexpensive, sensitive, and reliabl... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4273922 Wed, 01 Dec 2010 00:00:00 +0100 4273922 http://www.medworm.com/index.php?rid=4273921&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21143210%26dopt%3DAbstract Authors: Tanaka Y, Kimura Y, Mitani Y, Kawai Y, Lezhava A, Noma S, Tagami M, Kawai J, Hayashizaki Y, Usui K In DNA amplification, the initial step of copying a target sequence from the template DNA-the so-called intermediate product generation step-is very important. In examining the turn-back primer (TP)-dependent isothermal DNA amplification (TIA) method, we determined the actual time point of intermediate product generation by extrapolating dsDNA amplification curves. Our results indicate that intermediate product creation is the rate-limiting step in TIA, and good TP design is advantageous for improving the intermediate production process. Address correspondence to Kengo Usui. RIKEN Omics Science Center (OSC), RIKEN Yokohama Institute, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kana... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4273921 Wed, 01 Dec 2010 00:00:00 +0100 4273921 http://www.medworm.com/index.php?rid=4273920&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21143211%26dopt%3DAbstract Authors: Brisco M, Latham S, Bartley P, Morley A Optimal accuracy of quantitative PCR (qPCR) requires correction for integrity of the target sequence. Here we combine the mathematics of the Poisson distribution and exponential amplification to show that the frequency of lesions per base (which prevent PCR amplification) can be derived from the slope of the regression line between cycle threshold (Ct) and amplicon length. We found that the amplifiable fraction (AF) of a target can be determined from this frequency and the target length. Experimental results from this method correlated with both the magnitude of a damaging agent and with other measures of DNA damage. Applying the method to a reference sequence, we determined the values for lesions/base in control samples, as well as in t... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4273920 Wed, 01 Dec 2010 00:00:00 +0100 4273920 http://www.medworm.com/index.php?rid=4273919&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21143212%26dopt%3DAbstract Authors: Sengupta S, Ruotti V, Bolin J, Elwell A, Hernandez A, Thomson J, Stewart R Preparation of an Illumina sequencing library for gene expression analysis (mRNA-Seq) requires microgram amounts of starting total RNA or PCR-based amplification. Here we describe a protocol based on T7 linear RNA amplification that does not introduce significant bias, requires only 10 ng total RNA, and generates a directional, fully representative, whole-transcript mRNA-Seq Illumina library that is highly consistent across over three orders of magnitude of input RNA. Address correspondence to Ron Stewart, Regenerative Biology Laboratory, Morgridge Institute for Research, Genetics-Biotechnology Center Building, 425 Henry Mall, Madison, WI 53706,USA. e-mail: PMID: 21143212 [PubMed - in process] (Sour... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4273919 Wed, 01 Dec 2010 00:00:00 +0100 4273919 http://www.medworm.com/index.php?rid=4216586&cid=s_36935_70_f&fid=36935&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21091444%26dopt%3DAbstract Authors: Kodama Y, Hu CD Protein-protein interactions (PPIs) play crucial roles in various biological processes. Among biochemical, genetic, and imaging approaches that have been used for the study of PPIs, visualization of PPIs in living cells is the key to understanding their cellular functions. The bimolecular fluorescence complementation (BiFC) assay represents one of these imaging tools for direct visualization of PPIs in living cells. The BiFC assay is based on the structural complementation of two nonfluorescent N- and C-terminal fragments of a fluorescent protein when they are fused to a pair of interacting proteins. Although over 10 different fluorescent proteins have been used for BiFC assays, the two nonfluorescent fragments from all of these fluorescent proteins can spontan... BioTechniques journals http://www.medworm.com/rss/comments.php?id=4216586 Mon, 01 Nov 2010 00:00:00 +0100 4216586