BMC Biotechnology - Latest articles via MedWorm.com

Replication methods and tools in high-throughput cultivation processes - recognizing potential variations of growth and product formation by on-line monitoring

BMC Biotechnology - Latest articles — Tue, 16 Mar 2010 00:00:00 +0100

Conclusions: To improve the reproducibility of high-throughput cultivation processes and the comparability between different applied cultures, it is strongly recommended to use automated or manual liquid handling stations or, alternatively, multi-channel pipettes. Because of their higher transfer volume and hence precision in comparison to pin replicators, they reduce the variance of initial biomass concentrations. With respect to the results obtained, other methods to increase the comparability between parallel cultivations by compensating differences in biomass concentrations are required, such as using autoinduction media, fed-batch operation of precultures or on-line monitoring in microtiter plates combined with automated liquid handling. (Source: BMC Biotechnology - Latest articles)

A mutant Pfu DNA polymerase designed for advanced uracil-excision DNA engineering

BMC Biotechnology - Latest articles — Tue, 16 Mar 2010 00:00:00 +0100

Conclusions: The different uracil-excision based molecular tools that have been developed in an open-source fashion, constitute a comprehensive, yet simple and inexpensive toolkit for any need in molecular cloning. (Source: BMC Biotechnology - Latest articles)

Heterologous expression of a tannic acid-inducible laccase3 of Cryphonectria parasitica in Saccharomyces cerevisiae

BMC Biotechnology - Latest articles — Wed, 24 Feb 2010 00:00:00 +0100

Conclusions: The results suggest that expression of the lac3 gene has an inhibitory effect on the growth of transformed S. cerevisiae and that the controlled expression of lac3 is appropriate for the possible application of recombinant yeast to the treatment of phenolic compounds. (Source: BMC Biotechnology - Latest articles)

PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the Arabidopsis genome

BMC Biotechnology - Latest articles — Tue, 23 Feb 2010 00:00:00 +0100

Conclusion: The phiC31 system performs site-specific recombination in germinaltissue, a prerequisite for generating stable lines with unwanted DNA removed. Theprecise site-specific deletion by phiC31 in planta demonstrates that the recombinase can be used to remove selectable markers or other introduced transgenes that are no longer desired and therefore can be a useful tool for genome engineering in plants. (Source: BMC Biotechnology - Latest articles)

Immunohistochemical detection of transgene expression in the brain using small epitope tags

BMC Biotechnology - Latest articles — Thu, 18 Feb 2010 00:00:00 +0100

Conclusions: We show here that several small epitope tags are useful for immunohistochemical detection of exogenous proteins in vivo. Our study also provides a generic methodology which is broadly applicable for the detection of overexpressed transgenes in mammalian brain tissue. (Source: BMC Biotechnology - Latest articles)

Characterization of a molecular switch system that regulates gene expression in mammalian cells through a small molecule

BMC Biotechnology - Latest articles — Thu, 18 Feb 2010 00:00:00 +0100

Conclusions: We have successfully shown that this system can induce tightly controlled transgene expression and can be used for transient transfections or retroviral transductions in mammalian cell culture. Further characterization is needed for gene therapy applications. (Source: BMC Biotechnology - Latest articles)

High-level expression and purification of soluble recombinant FGF21 protein by SUMO fusion in Escherichia coli

BMC Biotechnology - Latest articles — Wed, 17 Feb 2010 00:00:00 +0100

Background: Fibroblast growth factor 21 (FGF21) is a promising drug candidate to combat metabolic diseases. However, high-level expression and purification of recombinant FGF21 (rFGF21) in Escherichia coli (E. coli) is difficult because rFGF21 forms inclusion bodies in the bacteria making it difficult to purify and obtain high concentrations of bioactive rFGF21. To overcome this problem, we fused the FGF21 with SUMO (Small ubiquitin-related modifier) by polymerase chain reaction (PCR), and expressed the fused gene in E.coli BL21(DE3). Results: By inducing with IPTG, SUMO-FGF21 was expressed at a high level. Its concentration reached 30% of total protein, and exceeded 95% of all soluble proteins. The fused protein was purified by DEAE sepharose FF and Ni-NTA affinity chromatography. Once cl...

Protein microarray: sensitive and effective immunodetection for drug residues

BMC Biotechnology - Latest articles — Tue, 16 Feb 2010 00:00:00 +0100

Background: Veterinary drugs such as clenbuterol (CL) and sulfamethazine (SM2) are low molecular weight ( (Source: BMC Biotechnology - Latest articles)

High-throughput detection of mutations responsible for childhood hearing loss using resequencing microarrays

BMC Biotechnology - Latest articles — Wed, 10 Feb 2010 00:00:00 +0100

Conclusions: Together, these findings provide insight into critical issues for disease-centered resequencing protocols suitable for clinical application and support the use of array-based resequencing technology as a valuable molecular diagnostic tool for pediatric SNHL and other genetic diseases with substantial genetic heterogeneity. (Source: BMC Biotechnology - Latest articles)

A versatile polyacrylamide gel electrophoresis based sulfotransferase assay

BMC Biotechnology - Latest articles — Wed, 10 Feb 2010 00:00:00 +0100

Conclusion: The versatility of the current method comes from that SDS-PAGE can separate proteins and small molecules according to different parameters. While mobilities of proteins during SDS-PAGE are inversely related to their sizes, mobilities of small molecules are positively related to their charge/mass ratios. The predicted relative mobility of a product to PAPS is a good indicator of whether a sulfotransferase can be assayed using SDS-PAGE. Because phosphorylation is most similar to sulfation in chemistry, the method is likely to be applicable to kinases as well. (Source: BMC Biotechnology - Latest articles)

Comparative transfection of DNA into primary and transformed mammalian cells from different lineages

BMC Biotechnology - Latest articles — Mon, 08 Feb 2010 00:00:00 +0100

Conclusions: In general, the Amaxa electroporation/nucleofection system appears to superior to other chemical systems. However, there are cell-type and species specific differences that need to be evaluated empirically to optimize the conditions for transfection efficiency and cell survival. (Source: BMC Biotechnology - Latest articles)

Periplasmic expression of soluble single chain T cell receptors is rescued by the chaperone FkpA

BMC Biotechnology - Latest articles — Wed, 03 Feb 2010 00:00:00 +0100

Conclusion: The general expression regime presented here allows for rapid production of soluble scTCRs and is applicable for 1) high yield recovery sufficient for biophysical characterization and 2) high throughput screening of such molecules following molecular engineering. (Source: BMC Biotechnology - Latest articles)

Heterologous expression of leader-less pga gene in Pichia pastoris: intracellular production of prokaryotic enzyme

BMC Biotechnology - Latest articles — Wed, 03 Feb 2010 00:00:00 +0100

Conclusions: Presented results provide useful data regarding fermentation strategy, intracellular biosynthetic potential, and consequences of the heterologous expression of PGAEc in P. pastoris X-33. Aberrant processing of the precursor of PGAEc in the cytosol yielded the mature enzyme with modified traits. (Source: BMC Biotechnology - Latest articles)

A new approach to dual-color two-photon microscopy with fluorescent proteins

BMC Biotechnology - Latest articles — Tue, 02 Feb 2010 00:00:00 +0100

Conclusion: Our method does not require large differences in Stokes shifts and can be extended to a variety of FP pairs with larger 2PA efficiency and more optimal imaging properties. (Source: BMC Biotechnology - Latest articles)

Senescent vs. non-senescent cells in the human annulus in vivo: Cell harvest with laser capture microdissection and gene expression studies with microarray analysis

BMC Biotechnology - Latest articles — Thu, 28 Jan 2010 00:00:00 +0100

Conclusions: Our LCM-microarray analyses identified a set of genes associated with senescence which were significantly upregulated in senescent vs non-senescent cells in the human annulus. These genes include p38 MAP kinase, discoidin, inhibitor of growth family member 5, and growth arrest and DNA-damage-inducible beta. Other genes, including genes associated with cell proliferation, extracellular matrix formation, cell signaling and other cell functions also showed significant modulation in senescent vs non-senescent cells. The aging/degenerating disc undergoes a well-recognized loss of cells; understanding senescent cells is important since their presence further reduces the disc's ability to generate new cells to replace those lost to necrosis or apoptosis. (Source: BMC Biotechnology - ...

A novel FRET pair for detection of parallel DNA triplexes by the LightCycler

BMC Biotechnology - Latest articles — Wed, 27 Jan 2010 00:00:00 +0100

Conclusions: We have established a novel pH independent FRET pair with high fluorescence signals on the LightCycler platform for both antiparallel duplex and parallel triplex formation. The method has been thoroughly validated, and is characterized by an excellent accuracy and reproducibility. This FRET pair is especially suitable for DeltaTm and Tm determinations of pH dependent parallel triplex formation. (Source: BMC Biotechnology - Latest articles)

Real-time monitoring of circadian clock oscillations in primary cultures of mammalian cells using Tol2 transposon-mediated gene transfer strategy

BMC Biotechnology - Latest articles — Fri, 22 Jan 2010 00:00:00 +0100

Conclusions: Our results indicate that gene transfer strategy using the Tol2 transposon system of a useful and safe non-viral vector is a powerful tool for investigating circadian rhythms in peripheral tissues. (Source: BMC Biotechnology - Latest articles)

Characterization of Proton Production and Consumption Associated with Microbial Metabolism

BMC Biotechnology - Latest articles — Wed, 20 Jan 2010 00:00:00 +0100

Conclusions: In this study, we have also shown that the final pH of the medium can be either acidic or basic depending on the choice of the electron acceptor for the same electron donor. In all cases, the in silico model could predict qualitatively the proton production/consumption rates obtained from the experimental data. Therefore, measurements of pH equivalents generated or consumed during growth can help characterize the microbial physiology further and can be valuable for optimizing practical applications such as microbial fuel cells, where growth associated pH changes can limit current generation rates. (Source: BMC Biotechnology - Latest articles)

Functional genomics tool: Gene silencing in Ixodes scapularis eggs and nymphs by electroporated dsRNA

BMC Biotechnology - Latest articles — Thu, 14 Jan 2010 00:00:00 +0100

Conclusions: Our study demonstrates that electroporation can be used as a simple dsRNA delivery tool in assessing the functional role of tick genes in the vector-host interactions. This technique represents a novel approach for specific gene suppression in immature stages of ticks. (Source: BMC Biotechnology - Latest articles)

RNA degradation compromises the reliability of microRNA expression profiling

BMC Biotechnology - Latest articles — Mon, 21 Dec 2009 00:00:00 +0100

Conclusion: MicroRNA expression cannot be reliably profiled in degraded total RNA. For the profiling of microRNAs we recommend use of RNA samples with a RNA integrity number equal to or above seven. (Source: BMC Biotechnology - Latest articles)

Unrestricted somatic stem cells from human umbilical cord blood grow in serum-free medium as spheres

BMC Biotechnology - Latest articles — Tue, 15 Dec 2009 00:00:00 +0100

Conclusions: These data suggest that USSC growth medium ACF maintains USSCs in an undifferentiated state and supports growth in suspension. This is the first demonstration that USSCs can grow in a serum- and animal component-free medium and that USSCs can form spheres. (Source: BMC Biotechnology - Latest articles)

An improved phage-display panning method to produce an HM-1 killer toxin anti-idiotypic antibody

BMC Biotechnology - Latest articles — Mon, 14 Dec 2009 00:00:00 +0100

Conclusion: The scFv antibodies generated after modified subtractive panning appear to have superior binding properties and cytocidal activity than regular panning. A simple modification of the elution condition in the phage-display panning protocol makes a large difference in determining success. Our method offers an attractive platform to discover potential therapeutic candidates. (Source: BMC Biotechnology - Latest articles)

Recombinant protein expression by targeting pre-selected chromosomal loci

BMC Biotechnology - Latest articles — Mon, 14 Dec 2009 00:00:00 +0100

Conclusion: RMCE provides a powerful method to specifically design gene expression with high accuracy. Upon considering the specific requirements of chromosomal sites this method provides a unique tool to exploit such sites for predictable expression of biotechnologically relevant proteins such as antibodies. (Source: BMC Biotechnology - Latest articles)

Jellyfish mucin may have potential disease modifying effects of osteoarthritis

BMC Biotechnology - Latest articles — Tue, 08 Dec 2009 00:00:00 +0100

Conclusion: At the concentrations of qniumucin used in this study, its injection together with HA inhibited articular cartilage degeneration in this model of OA. (Source: BMC Biotechnology - Latest articles)

A rapid and inexpensive labeling method for microarray gene expression analysis

BMC Biotechnology - Latest articles — Wed, 25 Nov 2009 00:00:00 +0100

Conclusion: The new direct random-primed cDNA labeling method introduced here is suitable for gene expression microarrays and provides a rapid, inexpensive alternative to existing methods. Using NimbleGen microarrays, the method produced excellent results comparable to those obtained with other methods. However, the simplicity and cost-effectiveness of the new method allows for increased sample throughput in microarray experiments and makes the process amenable to automation with a relatively simple liquid handling system. (Source: BMC Biotechnology - Latest articles)

Quantifying and resolving multiple vector transformants in S. cerevisiae plasmid libraries

BMC Biotechnology - Latest articles — Fri, 20 Nov 2009 00:00:00 +0100

Conclusions: These results highlight the potential for multiple vector transformants to dominate yeast libraries constructed by homologous recombination. The previously unrecognized prevalence and persistence of multiply transformed yeast cells have important implications for yeast library screens. The quantitative information described herein should increase awareness of this issue, and the rapid sequencing approach developed for these studies should be widely useful for identifying multiple vector transformants and avoiding complications associated with cells that have acquired more than one unique plasmid. (Source: BMC Biotechnology - Latest articles)

High-level HIV-1 Nef transient expression in Nicotiana benthamiana using the P19 gene silencing suppressor protein of Artichoke Mottled Crinckle Virus

BMC Biotechnology - Latest articles — Fri, 20 Nov 2009 00:00:00 +0100

Conclusions: We demonstrated that expression level of HIV-1 Nef in plant can be improved using a transient expression system enhanced by the AMCV-P19 gene silencing suppressor protein. Moreover, plant-derived Nef was purified, with enhanced yield, exploiting a two-step purification protocol. These results represent a first step towards the development of a plant-derived HIV vaccine. (Source: BMC Biotechnology - Latest articles)

Non-antibiotic selection systems for soybean somatic embryos: the lysine analog aminoethyl-cysteine as a selection agent

BMC Biotechnology - Latest articles — Wed, 18 Nov 2009 00:00:00 +0100

Conclusions: Genetically engineered soybeans expressing a lysine insensitive DHPS gene can be selected with the non-antibiotic selection agent AEC. We also report here the inhibitory effects of glufosinate, (isopropylamine-glyphosate) (Roundup(R)), AEC and the ALS inhibitors Exceed(R) and Synchrony(R) against different tissues of soybean. (Source: BMC Biotechnology - Latest articles)

Affinity maturation generates greatly improved xyloglucan-specific carbohydrate binding modules

BMC Biotechnology - Latest articles — Sat, 31 Oct 2009 00:00:00 +0100

Conclusions: We conclude that affinity maturation of CBM selected from molecular libraries based on the CBM4-2 scaffold is possible and has the potential to generate new analytical tools for detection of plant carbohydrates. (Source: BMC Biotechnology - Latest articles)

Genetic load and transgenic mitigating genes in transgenic Brassica rapa (field mustard) x Brassica napus (oilseed rape) hybrid populations

BMC Biotechnology - Latest articles — Sat, 31 Oct 2009 00:00:00 +0100

Conclusion: The effects of genetic load of crop and in some situations, weed alleles might be beneficial under certain environmental conditions. However, when genetic load was directly incorporated into transgenic events, e.g., using a TM construct, the number of transgenic hybrids and persistence in weedy genomic backgrounds was significantly decreased. (Source: BMC Biotechnology - Latest articles)

Novel immortalized human fetal liver cell line, cBAL111, has the potential to differentiate into functional hepatocytes

BMC Biotechnology - Latest articles — Tue, 20 Oct 2009 23:00:00 +0100

Conclusion: This novel liver cell line has the potential to differentiate into mature hepatocytes to be used for in vitro hepatocyte applications. (Source: BMC Biotechnology - Latest articles)

Characterization of unknown genetic modifications using high throughput sequencing and computational subtraction

BMC Biotechnology - Latest articles — Wed, 07 Oct 2009 23:00:00 +0100

Conclusion: We believe that computational subtraction represents a powerful new strategy for determining if an organism has been genetically modified as well as to define the nature of the modification. Fewer assumptions have to be made compared to methods currently in use and this is an advantage particularly when working with unknown GMOs. (Source: BMC Biotechnology - Latest articles)

Refinement of lentiviral vector for improved RNA processing and reduced rates of self inactivation repair

BMC Biotechnology - Latest articles — Tue, 06 Oct 2009 23:00:00 +0100

Conclusions: RNA processing can be improved by mutation of the major and minor HIV-1 splice donor sites in the vector. In addition the rate of vector mobilisation and repair of SIN vectors can be successfully reduced by careful vector design that reduces homology between the 5' and 3' long terminal repeats (LTRs) to a minimum. (Source: BMC Biotechnology - Latest articles)

Reduction of liver macrophage transduction by pseudotyping lentiviral vectors with a fusion envelope from Autographa californica GP64 and Sendai virus F2 domain.

BMC Biotechnology - Latest articles — Tue, 06 Oct 2009 23:00:00 +0100

Conclusions: We demonstrate reduced macrophage transduction in vitro and in vivo with GP64/Sendai chimeric envelope proteins. (Source: BMC Biotechnology - Latest articles)

Thermochemotherapy effect of nanosized As2O3/Fe3O4 complex on experimental mouse tumors and its influence on the expression of CD44v6, VEGF-C and MMP-9

BMC Biotechnology - Latest articles — Sun, 04 Oct 2009 23:00:00 +0100

Background: Both thermotherapy and arsenic have been shown to be active against a broad spectrum of cancers. To reduce the limitations of conventional thermotherapy, improve therapeutic anticancer activity, reduce the toxicity of arsenic on normal tissue, and increase tissue-specific delivery, we prepared a nanosized As2O3/Fe3O4 complex (Fe3O4 magnetic nanoparticles encapsulated in As2O3). We assessed the thermodynamic characteristics of this complex and validated the hyperthermia effect, when combined with magnetic fluid hyperthermia (MFH), on xenograft HeLa cells (human cervical cancer cell line) in nude mice. We also measured the effect on the expression of CD44v6, VEGF-C, and MMP-9 which were related to cancer and/or metastasis. Results: The nanosized As2O3/Fe3O4 particles were approxi...

Integrating human stem cell expansion and neuronal differentiation in bioreactors

BMC Biotechnology - Latest articles — Mon, 21 Sep 2009 23:00:00 +0100

Conclusions: The culture systems developed herein are robust and represent one-step-forward towards the development of integrated bioprocesses, bridging stem cell expansion and differentiation in fully controlled bioreactors. (Source: BMC Biotechnology - Latest articles)

A combination of transposable elements and magnetic cell sorting provides a very efficient transgenesis system for chicken primary erythroid progenitors

BMC Biotechnology - Latest articles — Thu, 17 Sep 2009 23:00:00 +0100

Conclusions: This new Tol2/MACS system leads to stable expression in a culture of primary chicken erythroid cells highly enriched in cells expressing the transgene of interest. This system could be used in a wide variety of vertebrate species. (Source: BMC Biotechnology - Latest articles)

Antibody expressing pea seeds as fodder for prevention of gastrointestinal parasitic infections in chickens

BMC Biotechnology - Latest articles — Thu, 10 Sep 2009 23:00:00 +0100

Conclusion: The results suggest that our strategy offers a general approach to control parasitic infections in production animals using cost-effective antibody expression in crop seeds affordable for the animal health market. (Source: BMC Biotechnology - Latest articles)

Monodispersity of recombinant Cre recombinase correlates with its effectiveness in vivo

BMC Biotechnology - Latest articles — Thu, 10 Sep 2009 23:00:00 +0100

Conclusions: A rational purification protocol for TAT-Cre has been developed by choosing the options that minimize the enzyme aggregation. Our data suggest that AI measurement should support the optimization of any protocol aiming at the recovery of monodispersed protein. (Source: BMC Biotechnology - Latest articles)

New stably transfected bioluminescent cells expressing FLAG epitope-tagged estrogen receptors to study their chromatin recruitment

BMC Biotechnology - Latest articles — Tue, 08 Sep 2009 23:00:00 +0100

Conclusion: In HELN derived cell lines, no fundamental differences between kinetics were observed during 8 hours for FLAG-ERalpha and FLAG-ERbeta, as well as for polymerase II recruitment. However, the relative importance of recruitment between 1 hr and 4 hr was found to be different in HeLa cell line expressing exogenous tagged ERalpha and in MCF-7 cell line expressing endogenous ER. (Source: BMC Biotechnology - Latest articles)

A comparative study of recombinant and native frutalin binding to human prostate tissues

BMC Biotechnology - Latest articles — Tue, 08 Sep 2009 23:00:00 +0100

Conclusions: Native and recombinant frutalin yielded different binding responses in the prostate tissues due to their differences in carbohydrate-binding affinities. Also, this study shows that both lectins may be used as histochemical biomarkers for the prostate cancer. Moreover, the successful use of a recombinant lectin in immunohistochemical studies of prostate cancer was for the first time demonstrated, highlighting the advantages of using recombinant systems in the preparation of pure lectin samples for diagnostic purpose. (Source: BMC Biotechnology - Latest articles)

Evaluation of the impact of single nucleotide polymorphisms and primer mismatches on quantitative PCR.

BMC Biotechnology - Latest articles — Thu, 27 Aug 2009 23:00:00 +0100

Conclusions: The ability to evaluate priming (and mispriming) rates and to predict their impacts provided a precise and quantitative description of assay performance. Priming probabilities were also found to be a good measure of analytical specificity. (Source: BMC Biotechnology - Latest articles)

Transduction of the MPG-tagged fusion protein into mammalian cells and oocytes depends on amiloride-sensitive endocytic pathway

BMC Biotechnology - Latest articles — Tue, 25 Aug 2009 23:00:00 +0100

Conclusions: The results show that the transduction of MPG fusion protein utilizes endocytic pathway(s) which is amiloride-sensitive and partially dynamin-dependent. Collectively, the MPG fusion protein could be further developed as a novel tool of "protein therapeutics", with potentials to be used in various cell systems including mammalian oocytes. (Source: BMC Biotechnology - Latest articles)

Directed evolution of a filamentous fungus for thermotolerance

BMC Biotechnology - Latest articles — Tue, 25 Aug 2009 23:00:00 +0100

Conclusion: We report the experimental evolution of a filamentous fungus via the novel application of a powerful new continuous culture device. This is the first example of using continuous culture to select for complex phenotypes such as thermotolerance. Temperature adapted variants of the insect-pathogenic, filamentous fungus M. anisopliae were isolated and demonstrated to show vigorous growth at a temperature that is inhibitory for the parent strain. Insect virulence assays confirmed that pathogenicity can be retained during the selection process. In principle, this technology can be used to adapt filamentous fungi to virtually any environmental condition including abiotic stress and growth substrate utilization. (Source: BMC Biotechnology - Latest articles)

High quality protein microarray using in situ protein purification

BMC Biotechnology - Latest articles — Sat, 22 Aug 2009 23:00:00 +0100

Conclusion: An optimized platform for in situ protein purification on microarray slides using His-tagged recombinant proteins is a desirable tool for the screening of novel protein functions and protein-protein interactions. In the context of immunoproteomics, such protein microarrays are complimentary to approaches using non-recombinant methods to discover and characterize bacterial antigens. (Source: BMC Biotechnology - Latest articles)

Archaeosomes made of Halorubrum tebenquichensetotal polar lipids: a new source of adjuvancy

BMC Biotechnology - Latest articles — Wed, 12 Aug 2009 23:00:00 +0100

Conclusion: We herein report the finding of new H. tebenquichense non alkaliphilic strains in Argentinean Patagonia together with the adjuvant properties of ARC after sc administration in mice. Our results indicate that archaeosomes prepared with TPL from these two strains could be successfully used as vaccine delivery vehicles. (Source: BMC Biotechnology - Latest articles)

Archaeosomes made of Halorubrum tebenquichense total polar lipids: A new source of adjuvancy

BMC Biotechnology - Latest articles — Wed, 12 Aug 2009 23:00:00 +0100

Conclusions: We herein report the finding of new H. tebenquichense non alkaliphilic strains in Argentinean Patagonia together with the adjuvant properties of ARC after sc administration in mice. Our results indicate that archaeosomes prepared with TPL from these two strains could be successfully used as vaccine delivery vehicles. (Source: BMC Biotechnology - Latest articles)

Efficient generation of long-distance conditional alleles using recombineering and a dual selection strategy in replicate plates

BMC Biotechnology - Latest articles — Mon, 27 Jul 2009 23:00:00 +0100

Conclusions: The strategy of ES cell cultures in replicate plates proved to be very efficient in identifying ES cells that had undergone the correct recombination event. This approach facilitates the generation of conditional knock-out mice when large parts of the genome are intended to be flanked by loxP sites. (Source: BMC Biotechnology - Latest articles)

Evaluation of cross-linked aggregates from purified Bacillus subtilis levansucrase mutants for transfructosylation reactions

BMC Biotechnology - Latest articles — Sun, 26 Jul 2009 23:00:00 +0100

Conclusions: This is the first report of cross-linked transglycosidases aggregates. CLEAs prepared from purified LS and mutants have the highest specific activity for immobilized fructosyltransferases (FTFs) reported in the literature. CLEAs from R360K and Y429N LS mutants were particularly suitable for fructosyl-xyloside synthesis as the absence of levan synthesis decreases diffusion limitation and increases operational stability. (Source: BMC Biotechnology - Latest articles)

Gamma-phage lysin PlyG sequence-based synthetic peptides coupled with Qdot-nanocrystals are useful for developing detection methods for Bacillus anthracis by using its surrogates, B. anthracis-Sterne and B. cereus-4342

BMC Biotechnology - Latest articles — Tue, 21 Jul 2009 23:00:00 +0100

Conclusion: Overall, these studies illustrate that the short 10-amino acid sequence 'LKMTADFILQ' in fact is a stand-alone bacterial cell wall-binding motif of PlyG. In principle, synthetic peptides PlyG-P1, PlyG-P3 and PlyG-P5, especially PlyG-P3 coupled with Qdot-nanocrystals are useful as high-sensitivity bio-probes in developing detection technologies for B. anthracis. (Source: BMC Biotechnology - Latest articles)

Efficient, chemoselective synthesis of immunomicelles using single-domain antibodies with a C-terminal thioester

BMC Biotechnology - Latest articles — Sun, 19 Jul 2009 23:00:00 +0100

Conclusion: A novel procedure was developed to obtain soluble, well-folded single-domain antibodies with reactive C-terminal thioesters in good yields. These proteins are promising building blocks for the chemoselective functionalization via NCL of a broad range of nanoparticle scaffolds, including micelles, liposomes and dendrimers. (Source: BMC Biotechnology - Latest articles)

A novel Poly(epsilon-caprolactone)-Pluronic-Poly(epsilon-caprolactone) grafted Polyethyleneimine(PCFC-g-PEI), Part 1, synthesis, cytotoxicity, and in vitro transfection study

BMC Biotechnology - Latest articles — Thu, 16 Jul 2009 23:00:00 +0100

Conclusion: The cytotoxicity and in vitro transfection study indicated that PCFC-g-PEI copolymer prepared in this paper was a novel gene delivery system with lower cytotoxicity and considerable transfection efficiency compared with commercial PEI (25kD). (Source: BMC Biotechnology - Latest articles)

Stable nuclear transformation of Gonium pectorale

BMC Biotechnology - Latest articles — Thu, 09 Jul 2009 23:00:00 +0100

Conclusion: Flanking sequences that include promoters from C. reinhardtii and from V. carteri work in G. pectorale and allow the functional expression of heterologous genes, such as the selectable marker gene aphVIII of S. rimosus or the co-transformed, codon-optimized G. princeps luciferase gene, which turned out to be a suitable reporter gene in Gonium. The availability of a method for transformation of Gonium makes genetic engineering of this species possible and allows for detailed studies in molecular evolution using the unicellular Chlamydomonas, the 16-celled Gonium, and the multicellular Volvox. (Source: BMC Biotechnology - Latest articles)

Production of N-acetyl-D-neuraminic Acid Using Two Sequential Enzymes Overexpressed as Double-Tagged Fusion Proteins

BMC Biotechnology - Latest articles — Wed, 08 Jul 2009 23:00:00 +0100

Conclusions: Double-tagged gene fusions were proposed for over-expressing two sequential enzyme proteins, which could be easily immobilzed onto ionic exchangers by direct capture from the crude prtoein extracts via ionic tags. The immobilized, double-tagged proteins were demonstrated to be effective for one-pot enzymatic production of sialic acid. (Source: BMC Biotechnology - Latest articles)

A further insight into the biosorption mechanism of Pt() by infrared spectrometry

BMC Biotechnology - Latest articles — Sun, 05 Jul 2009 23:00:00 +0100

Conclusion: That knowledge could serve as a guide in the researches for improving the preparation of highly dispersive supported platinum catalyst and for fabricating new advanced platinum nanostructured devices by biotechnological methods. (Source: BMC Biotechnology - Latest articles)

Isolation of a human-like antibody fragment (scFv) that neutralizes ricin biological activity

BMC Biotechnology - Latest articles — Mon, 29 Jun 2009 23:00:00 +0100

Conclusion: Results of the characterization studies, and the high degree of identity with human germline genes, altogether make this anti-ricin scFv, or an IgG derived from it, a likely candidate for use in humans to minimize effects caused by ricin intoxication. (Source: BMC Biotechnology - Latest articles)

Simple and efficient site-directed mutagenesis using two single-primer reactions in parallel to generate mutants for protein structure-function studies

BMC Biotechnology - Latest articles — Mon, 29 Jun 2009 23:00:00 +0100

Conclusion: The SPRINP mutagenesis protocol yields mutants reliably and with high fidelity. The use of a single primer in each amplification reaction increases the probability of success of primers relative to previous methods employing a forward and reverse primer pair in the same reaction. (Source: BMC Biotechnology - Latest articles)

Expression of endo-1, 4-beta-xylanase from Trichoderma reesei in Pichia pastoris and functional characterization of the produced enzyme

BMC Biotechnology - Latest articles — Mon, 15 Jun 2009 23:00:00 +0100

Conclusion: The P. pastoris expression system allows a high level expression of xylanases. Xylanase was the main protein species in the culture supernatant, and the functional tests indicated that even the non-purified enzyme shows highly specific xylanase activity that is free of cellulolytic side acitivities. Therefore, P pastoris is a very useful expression system when the goal is highly specific and large scale production of glycosyl hydrolases. (Source: BMC Biotechnology - Latest articles)

Interfering ribonucleic acids that suppress expression of multiple unrelated genes

BMC Biotechnology - Latest articles — Mon, 15 Jun 2009 23:00:00 +0100

Conclusions: Our work demonstrates the possibility of designing multitargeting dsRNA to suppress more than one disease-altering gene. This warrants further investigation as a possible therapeutic approach. (Source: BMC Biotechnology - Latest articles)

High-titer preparation of Bombyx mori nucleopolyhedrovirus (BmNPV) displaying recombinant protein in silkworm larvae by size exclusion chromatography and its characterization

BMC Biotechnology - Latest articles — Thu, 11 Jun 2009 23:00:00 +0100

Conclusion: The successful purification of rBmNPV-hPRR indicates that baculovirus production using silkworm larvae and its purification from hemolymph by Sephacryl S-1000 SF column chromatography can provide an economical approach in obtaining the purified BmNPV stocks with high titer for large-scale production of hPRR. Also, it can be utilized for further binding analysis and screening of inhibitors of hPRR. (Source: BMC Biotechnology - Latest articles)

Synthesis of sialoglycopolypeptide for potentially blocking Influenza virus infection using a rat alpha2,6-sialyltransferase expressed in BmNPV bacmid-injected silkworm larvae

BMC Biotechnology - Latest articles — Fri, 05 Jun 2009 04:00:00 +0100

Conclusions: The recombinant alpha2,6SiaT from a silkworm expression system is useful for the synthesis of sialylation of asialoglycopeptide and a valuable tool for investigating the molecular mechanisms of biological and physiological events, such as cell-cell recognition and viral infections. (Source: BMC Biotechnology - Latest articles)

Construction of siRNA/miRNA expression vectors based on a one-step PCR process

BMC Biotechnology - Latest articles — Tue, 02 Jun 2009 04:00:00 +0100

Background: RNA interference (RNAi) has become a powerful means for silencing target gene expression in mammalian cells and is envisioned to be useful in therapeutic approaches to human disease. In recent years, high-throughput, genome-wide screening of siRNA/miRNA libraries has emerged as a desirable approach. Current methods for constructing siRNA/miRNA expression vectors require the synthesis of long oligonucleotides, which is costly and suffers from mutation problems. Results: Here we report an ingenious method to solve traditional problems associated with construction of siRNA/miRNA expression vectors. We synthesized shorter primers ( (Source: BMC Biotechnology - Latest articles)

Integrated application of uniform design and least-squares support vector machines to transfection optimization

BMC Biotechnology - Latest articles — Sun, 31 May 2009 04:00:00 +0100

Conclusions: The integrated application of uniform design and least-squares support vector machine is a simple technique for obtaining high transfection efficiency. Using this novel method, the number of required experiments would be greatly cut down while higher efficiency would be gained. Least-squares support vector machine may be applicable to many other problems that need to be optimized. (Source: BMC Biotechnology - Latest articles)

A microarray-based method for the parallel analysis of genotypes and expression profiles of wood-forming tissues in Eucalyptus grandis

BMC Biotechnology - Latest articles — Wed, 27 May 2009 04:00:00 +0100

Conclusions: A simple, yet general method was developed for genotyping and expression profiling of wood-forming tissue of E. grandis trees differing in their splitting characteristics and in their lignin contents. Evaluation of gene expression profiles and the binary distribution of cDNA-AFLP fragments on the chip suggest that the prototype chip developed could be useful for transcript profiling and for the identification of Eucalyptus trees with preferred wood quality traits in commercial breeding programmes. (Source: BMC Biotechnology - Latest articles)

Expanding the multicolor capabilities of basic confocal microscopes by employing red and near-infrared quantum dot conjugates

BMC Biotechnology - Latest articles — Fri, 22 May 2009 04:00:00 +0100

Conclusion: Our data demonstrate that nearly any basic confocal microscope can be modified by the simple addition of appropriate emission filters, allowing the detection of red and near-infrared quantum dot conjugates. Additionally, quantum dot- and organic dye-based secondary reagents can be successfully combined in complex intracellular staining experiments. Substantial expansion of the multi-parameter capabilities of basic confocal instruments can be achieved with a financial investment that is minimal in comparison to instrument replacement or upgrade with additional lasers. (Source: BMC Biotechnology - Latest articles)

Cost-effective HRMA pre-sequence typing of clone libraries; application to phage display selection

BMC Biotechnology - Latest articles — Fri, 22 May 2009 04:00:00 +0100

Conclusions: Using HRMA, analysis of up to 384 samples can be done simultaneously and will take approximately 30 minutes. Clustering of clones can be largely automated using the system's software within 2 hours. Applied to the analysis of clones obtained after phage display antibody selection, HRMA facilitated a quick overview of the overall success as well as the identification of identical clones. Our approach can be used to characterize any clone set prior to sequencing, thereby reducing sequencing costs significantly. (Source: BMC Biotechnology - Latest articles)

Luciferase activity under direct ligand-dependent control of a muscarinic acetylcholine receptor

BMC Biotechnology - Latest articles — Mon, 18 May 2009 04:00:00 +0100

Conclusion: Our results demonstrate that in engineered GPCR-enzyme chimeras, intracellular enzyme activity can be directly controlled by a GPCR serving as the extracellular ligand-binding domain. (Source: BMC Biotechnology - Latest articles)

RNAi in the cereal weevil Sitophilus spp: Systemic gene knockdown in the bacteriome tissue

BMC Biotechnology - Latest articles — Fri, 15 May 2009 04:00:00 +0100

Conclusion: These results demonstrate that whole larval injection with dsRNA results in a high and systemic decrease of both mRNA and protein in the bacteriome tissue. This, along with the possibility of access to the insect developmental stages, opens up a new research avenue for exploring gene specific functions in the cereal weevils. (Source: BMC Biotechnology - Latest articles)

Fluorescent labeling of NASBA amplified tmRNA molecules for microarray applications

BMC Biotechnology - Latest articles — Fri, 15 May 2009 04:00:00 +0100

Conclusions: We have successfully demonstrated efficient combination of NASBA amplification technology with microarray based hybridization detection. Method is applicative for many different areas of microbial diagnostics including environmental monitoring, bio threat detection, industrial process monitoring and clinical microbiology. (Source: BMC Biotechnology - Latest articles)

The CD-loop of PAI-2 (SERPINB2) is redundant in the targeting, inhibition and clearance of cell surface uPA activity

BMC Biotechnology - Latest articles — Thu, 14 May 2009 04:00:00 +0100

Conclusions: We demonstrate that the CD-loop is redundant for the purposes of cellular uPA inhibition and cell surface clearance (endocytosis) and is thus suitable for the development of anti-uPA targeted cancer therapeutics. (Source: BMC Biotechnology - Latest articles)

Fluorescent labeling in semi-solid medium for selection of mammalian cells secreting high-levels of recombinant proteins

BMC Biotechnology - Latest articles — Mon, 11 May 2009 04:00:00 +0100

Conclusions: FLSSM allows rapid visualization of the high secretors from the transfected pools prior to picking, thus eliminating the tedious task of screening a high number of cell isolates. Because of its rapidity and its simplicity, FLSSM is a versatile method for the screening of high producers for research and industry. (Source: BMC Biotechnology - Latest articles)

A suite of recombinant luminescent bacterial strains for the quantification of bioavailable heavy metals and toxicity testing

BMC Biotechnology - Latest articles — Fri, 08 May 2009 04:00:00 +0100

Conclusions: The constructed battery of 19 luminescent sensor strains exhibits several novel aspects as it contains i) different bacterial hosts with similar metal-responsive elements ii) similar bacteria with metal-responsive elements in different copies in the cell and iii) a lights-off control for every metal sensor strain. No Gram-positive metal sensor expressing full bacterial bioluminescence system was constructed earlier. The combinations of different constructed strains enabled to follow the effect of the bacterial host on their sensitivity and specificity as well as on bioavailability of metals in various samples. (Source: BMC Biotechnology - Latest articles)

Efficient CPP -mediated Cre protein delivery to developing and adult CNS tissues.

BMC Biotechnology - Latest articles — Fri, 24 Apr 2009 04:00:00 +0100

Conclusions: We provide tools for the spatially restricted genetic modification of cells in explant culture. This strategy allows to study lineage, migration, differentiation and death of neural cells. As a proof-of-concept applied to CNS tissue, direct delivery of Cre recombinase enabled the selective elimination of an interneuron subpopulation of the spinal cord, thereby providing a model to study early events of neurodegenerative processes. Thus our work opens new perspectives for both fundamental and applied cell targeting protocols using proteic cargoes which need to retain full bioactivity upon internalisation, as illustrated here with the oligomeric Cre recombinase. (Source: BMC Biotechnology - Latest articles)

Acetylation of PAMAM dendrimers for cellular delivery of siRNA

BMC Biotechnology - Latest articles — Thu, 23 Apr 2009 04:00:00 +0100

Conclusions: Primary amine acetylation of PAMAM dendrimers reduced their cytotoxicity to U87 cells, and promoted the release of siRNA from dendrimer/siRNA complexes. A modest fraction (approximately 20%) of primary amines of PAMAM can be modified while maintaining the siRNA delivery efficiency of unmodified PAMAM, but higher degrees of amine neutralization reduced the gene silencing efficiency of PAMAM/siRNA delivery vectors. (Source: BMC Biotechnology - Latest articles)

A modified and automated version of the 'Fluorimetric Detection of Alkaline DNA Unwinding' method to quantify formation and repair of DNA strand breaks

BMC Biotechnology - Latest articles — Thu, 23 Apr 2009 04:00:00 +0100

Conclusion: We have established a robust and convenient method measuring of formation and repair of DNA single-strand breaks in live cells. While the sensitivity of our method is comparable to current assays, throughput is massively increased while operator time is decreased. (Source: BMC Biotechnology - Latest articles)

Gene Composer: database software for protein construct design, codon engineering, and gene synthesis

BMC Biotechnology - Latest articles — Tue, 21 Apr 2009 04:00:00 +0100

Conclusions: We present a complete description of Gene Composer functionality, and an efficient PCR-based synthetic gene assembly procedure with mis-match specific endonuclease error correction in combination with PIPE cloning. In a sister manuscript we present data on how Gene Composer designed genes and protein constructs can result in improved protein production for structural studies. (Source: BMC Biotechnology - Latest articles)

Combined protein construct and synthetic gene engineering for heterologous protein expression and crystallization using Gene Composer

BMC Biotechnology - Latest articles — Tue, 21 Apr 2009 04:00:00 +0100

Conclusion: The results consistently demonstrate that protein yields from codon engineered Gene Composer designs are as good as or better than those achieved from the synonymous native genes. Moreover, structure guided N- and C-terminal deletion constructs designed with the aid of Gene Composer can lead to greater success in gene to structure work as exemplified by the X-ray crystallographic structure determination of FtsZ from Bacillus subtilis. These results validate the Gene Composer algorithms, and suggest that using a combination of synthetic gene and protein construct engineering tools can improve the economics of gene to structure research. (Source: BMC Biotechnology - Latest articles)

Collagen matrices from sponge to nano: new perspectives for tissue engineering of skeletal muscle?

BMC Biotechnology - Latest articles — Wed, 15 Apr 2009 04:00:00 +0100

Conclusion: Collagen I-fibrin mixtures as well as collagen nanofibers yield good proliferation rates and myogenic differentiation of primary rat myoblasts in vitro. In addition parallel orientated nanofibers enable the generation of aligned cell layers and therefore represent the most promising step towards successful engineering of skeletal muscle tissue. (Source: BMC Biotechnology - Latest articles)

Noncytotoxic Orange and Red/Green Derivatives of DsRed-Express2 for Whole-Cell Labeling

BMC Biotechnology - Latest articles — Fri, 03 Apr 2009 04:00:00 +0100

Conclusion: E2-Orange and E2-Red/Green will facilitate the production of healthy, stably fluorescent cell lines and transgenic organisms for multi-color labeling studies. (Source: BMC Biotechnology - Latest articles)

Optimization of codon composition and regulatory elements for expression of human insulin like growth factor-1 in transgenic chloroplasts and evaluation of structural identity and function

BMC Biotechnology - Latest articles — Fri, 03 Apr 2009 04:00:00 +0100

Conclusions: This study demonstrates that the human Insulin like Growth Factor-1 expressed in transgenic chloroplasts is identical to the native protein and is fully functional. The ability to use plant chloroplasts as bioreactors to generate proteins of great economic value that retain their biological activity is an exciting and achievable goal that appears to be within our grasp. (Source: BMC Biotechnology - Latest articles)

Simultaneous detection of mRNA and protein stem cell markers in live cells

BMC Biotechnology - Latest articles — Thu, 02 Apr 2009 04:00:00 +0100

Conclusions: Molecular beacons have the potential to provide a powerful tool for highly specific detection and isolation of stem cells, including cancer stem cells and induced pluripotent stem (iPS) cells without disturbing cell physiology. It is advantageous to perform simultaneous detection of intracellular (mRNA) and cell-surface (protein) stem cell markers in flow cytometric analysis, which may lead to high detection sensitivity and efficiency. (Source: BMC Biotechnology - Latest articles)

PhiC31 integrase induces a DNA damage response and chromosomal rearrangements in human adult fibroblasts

BMC Biotechnology - Latest articles — Thu, 02 Apr 2009 04:00:00 +0100

Conclusions: In primary human fibroblasts expression of PhiC31 integrase leads to a DNA damage response and chromosomal aberrations. (Source: BMC Biotechnology - Latest articles)

Vaccination with recombinant Boophilus annulatus Bm86 ortholog protein, Ba86, protects cattle against B. annulatus and B. microplus infestations

BMC Biotechnology - Latest articles — Tue, 31 Mar 2009 04:00:00 +0100

Conclusion: These experiments showed the efficacy of recombinant Ba86 for the control of B. annulatus and B. microplus infestations in cattle and suggested that physiological differences between B. microplus and B. annulatus and those encoded in the sequence of Bm86 orthologs may be responsible for the differences in susceptibility of these tick species to Bm86 vaccines. (Source: BMC Biotechnology - Latest articles)

AC133+ progenitor cells as gene delivery vehicle and cellular probe in subcutaneous tumor models: a preliminary study

BMC Biotechnology - Latest articles — Fri, 27 Mar 2009 04:00:00 +0100

In this study, we used superparamagentic iron oxide (SPIO)-labeled APCs to carry the human sodium iodide symporter (hNIS) gene to the sites of implanted breast cancer in mouse model. In vivo real time tracking of these cells was performed by MRI and expression of hNIS was determined by Tc-99m pertechnetate (Tc-99m) scan. Results: Three million human breast cancer (MDA-MB-231) cells were subcutaneously implanted in the right flank of nude mice. APCs, isolated from fresh human cord blood, were genetically transformed to carry the hNIS gene using adenoviral vectors and magnetically labeled with ferumoxides-protamine sulfate (FePro) complexes. Magnetically labeled genetically transformed cells were administered intravenously in tumor bearing mice when tumors reached 0.5 cm in the largest dime...

Effects of research tool patents on biotechnology innovation in a developing country: A case study of South Korea

BMC Biotechnology - Latest articles — Thu, 26 Mar 2009 04:00:00 +0100

Conclusions: On the basis of the results of this investigation in combination with those of previous studies, we concluded that although restricted access problems have occurred, this has not yet deterred innovation in Korea. However, potential problems do exist, and the effects of restricted access should be constantly scrutinized. (Source: BMC Biotechnology - Latest articles)

Accumulation and processing of a recombinant protein designed as a cleavable fusion to the endogenous Rubisco LSU protein in Chlamydomonas chloroplast

BMC Biotechnology - Latest articles — Thu, 26 Mar 2009 04:00:00 +0100

Conclusion: These results demonstrate the utility of fusion proteins in algal chloroplast as a method to increase accumulation of recombinant proteins that are difficult to express. Since Rubisco is ubiquitous to land plants and green algae, this strategy may also be applied to higher plant transgenic expression systems. (Source: BMC Biotechnology - Latest articles)

Folding machineries displayed on a cation-exchanger for the concerted refolding of cysteine- or proline-rich proteins

BMC Biotechnology - Latest articles — Thu, 26 Mar 2009 04:00:00 +0100

Conclusions: The refolding-facilitating media presented here could be a cost-efficient platform and should be applicable to refold a wide range of E. coli inclusion bodies in high yield with biological function. (Source: BMC Biotechnology - Latest articles)

Practical and reliable FRET/FLIM pair of fluorescent proteins

BMC Biotechnology - Latest articles — Wed, 25 Mar 2009 04:00:00 +0100

Conclusions: The combined advantages suggest that the TagGFP-TagRFP is one of the most efficient green/red couples available to date for FRET/FLIM analyses to monitor interaction of proteins of interest in living cells and to generate FRET-based sensors for various applications. CaspeR3 provides reliable detection of apoptosis, and should become a popular tool both for cell biology studies and high throughput screening assays. (Source: BMC Biotechnology - Latest articles)

Viral and murine interleukin-10 are correctly processed and retain their biological activity when produced in tobacco

BMC Biotechnology - Latest articles — Thu, 19 Mar 2009 04:00:00 +0100

Conclusions: Tobacco plants are able to correctly process viral and murine IL-10 into biologically active dimers, therefore representing a suitable platform for the production for these cytokines. The accumulation levels obtained are high enough to allow delivery of an immunologically relevant dose of IL-10 in a reasonable amount of leaf material, without extensive purification. This study paves the way to performing feeding studies in mouse models of autoimmune diseases, that will allow the evaluation of the immunomodulatory properties and effectiveness of the viral IL-10 in inducing oral tolerance compared to the murine one. (Source: BMC Biotechnology - Latest articles)

Adenoviral vectors for highly selective gene expression in central serotonergic neurons reveal quantal characteristics of serotonin release in the rat brain

BMC Biotechnology - Latest articles — Thu, 19 Mar 2009 04:00:00 +0100

Conclusions: For the first time quantal release of 5HT from somato-dendritic compartments and axonal varicosities in mammalian brain has been demonstrated directly and characterised. Release from somato-dendritic and axonal compartments might have different physiological functions. The novel vectors generated in this study open a host of new experimental opportunities and will greatly facilitate further studies of the central serotonergic system. (Source: BMC Biotechnology - Latest articles)

A novel method for producing target cells and assessing cytotoxic T lymphocyte activity in outbred hosts

BMC Biotechnology - Latest articles — Wed, 11 Mar 2009 04:00:00 +0100

Conclusions: Given the versatility of the bicistronic vector used, its ability to deliver multiple and large transgenes in target cells, and its extremely wide cell specificity when pseudotyped with the vesicular stomatitis virus envelope protein, the method is potentially exploitable in many animal species. (Source: BMC Biotechnology - Latest articles)

An optimised system for refolding of human glucose 6-phosphate dehydrogenase

BMC Biotechnology - Latest articles — Wed, 11 Mar 2009 04:00:00 +0100

Conclusions: L-Arg is the key player in the refolding of human G6PD, preventing the aggregation of folding intermediate, and NADP+ is essential for the folding intermediate to adopt native structure. The refolding protocol can be applied to produce high recovery yield of folded protein with unaltered properties, paving the way for future studies on clinical G6PD mutants with folding defects and providing a useful model system to study the folding process of oligomeric proteins. (Source: BMC Biotechnology - Latest articles)

Addition of a sequence from α2-antiplasmin transforms human serum albumin into a blood clot component that speeds clot lysis

BMC Biotechnology - Latest articles — Tue, 03 Mar 2009 05:00:00 +0100

Conclusion: In this first report of transfer of transglutamination substrate status from one plasma protein to another, fusion protein α2AP(13-42)-HSA was shown to satisfy initial requirements for a long-lasting, well-tolerated competitive inhibitor of α2-antiplasmin predicted to act in a clot-localized manner. (Source: BMC Biotechnology - Latest articles)

Addition of a sequence from alpha-2-antiplasmin transforms human serum albumin into a blood clot component that speeds clot lysis

BMC Biotechnology - Latest articles — Tue, 03 Mar 2009 05:00:00 +0100

Conclusions: In this first report of transfer of transglutamination substrate status from one plasma protein to another, fusion protein alpha-2-AP(13-42)-HSA was shown to satisfy initial requirements for a long-lasting, well-tolerated competitive inhibitor of alpha-2-antiplasmin predicted to act in a clot-localized manner. (Source: BMC Biotechnology - Latest articles)

A multi-Fc-species system for recombinant antibody production

BMC Biotechnology - Latest articles — Thu, 26 Feb 2009 05:00:00 +0100

Conclusions: Altogether, this new expression system, that brings constant quality, sensitivity and unique versatility, will be important to broaden the use of recombinant and natural monoclonal antibodies both for laboratory and diagnosis use. (Source: BMC Biotechnology - Latest articles)

Incorporating double copies of a chromatin insulator into lentiviral vectors results in less viral integrants

BMC Biotechnology - Latest articles — Tue, 24 Feb 2009 05:00:00 +0100

Conclusion: These results have importance for the future use of insulator sequences in lentiviral vectors and might limit the use of insulators in vectors for in vivo use. Therefore, a careful analysis of the optimal design must be performed before insulators are included into clinical lentiviral vectors. (Source: BMC Biotechnology - Latest articles)

Improving the functional expression of a Bacillus licheniformis laccase by random and site-directed mutagenesis

BMC Biotechnology - Latest articles — Mon, 23 Feb 2009 05:00:00 +0100

Conclusions: The constructed double mutant K316N/D500G of the Bacillus licheniformis CotA laccase is an appropriate candidate for biotechnological applications due to its high expression level and high activity in dimerization of phenolic acids and decolorization of industrial dyes. (Source: BMC Biotechnology - Latest articles)

Noninvasive technique for measurement of heartbeat regularity in zebrafish (Danio rerio) embryos

BMC Biotechnology - Latest articles — Thu, 19 Feb 2009 05:00:00 +0100

Conclusion: The data support and validate this rapid, simple, noninvasive method, which includes video image analysis and frequency analysis. This method is capable of measuring the heart rate and heartbeat regularity simultaneously via the analysis of caudal blood flow in zebrafish embryos. With the advantages of rapid sample preparation procedures, automatic image analysis and data analysis, this method can potentially be applied to cardiotoxicity screening assay. (Source: BMC Biotechnology - Latest articles)

In vitro Drug Release Behavior from a Novel Thermosensitive Composite Hydrogel Based on Pluronic F127 and Poly(ethylene glycol)-Poly(epsilon-caprolactone)-Poly(ethylene glycol) Copolymer

BMC Biotechnology - Latest articles — Wed, 11 Feb 2009 05:00:00 +0100

Conclusions: The results showed that composite hydrogel prepared in this paper were biocompatible with low cell cytotoxicity, and the drugs in this work could be released slowly from composite hydrogel in an extended period, which suggested that the composite hydrogel might have great potential applications in biomedical fields. (Source: BMC Biotechnology - Latest articles)

A compact phage display human scFv library for selection of antibodies to a wide variety of antigens

BMC Biotechnology - Latest articles — Thu, 29 Jan 2009 05:00:00 +0100

Conclusions: These results suggested that by using an optimized protocol and very high repertoire diversity, a compact and efficient phage antibody library can be generated. This advanced method could be adopted by any molecular biology laboratory to generate both naive or immunized libraries for particular targets as well as for high-throughput applications. (Source: BMC Biotechnology - Latest articles)

Albumin fibrillization induces apoptosis via integrin/FAK/Akt pathway

BMC Biotechnology - Latest articles — Thu, 08 Jan 2009 05:00:00 +0100

Conclusions: We report on a novel process for converting globular proteins into fibrillar form to cause apoptosis by modulating the integrin/FAK/Akt/GSK-3beta/caspase-3 signaling pathway. Our findings may be useful for understanding the pathogenesis of amyloid-like fibrils and applicable for the development of better therapeutic agents that target the underlying mechanism(s) of the etiologic agents. (Source: BMC Biotechnology - Latest articles)

Single-chain Fv phage display propensity exhibits strong positive correlation with overall expression levels

BMC Biotechnology - Latest articles — Mon, 29 Dec 2008 05:00:00 +0100

Background: Single chain Fvs (scFvs) are widely applied in research, diagnostics and therapeutic settings. Display and selection from combinatorial libraries is the main route to their discovery and many factors influence the success of this process. They exhibit low thermodynamic stability, resulting in low levels of premature cytosolic folding or aggregation and are therefore compatible with post-translational translocation via SecYEG and display on phage in E. coli. However, there is little information indicating how this is related to bacteriophage display. Results: We characterised the relationship between overall scFv expression and display propensity for a panel of 15 anti-tetanus toxin scFvs and found a strong positive correlation (Rho=0.88, p (Source: BMC Biotechnology - Latest ar...

Better estimation of protein-DNA interaction parameters improve prediction of functional sites

BMC Biotechnology - Latest articles — Tue, 23 Dec 2008 05:00:00 +0100

Conclusions: We think that training binding site detection algorithms on datasets from binding assays lead to better prediction. The improvements in accuracy came from the unbiased nature of the SELEX dataset rather than from the number of sites available. We believe that with progress in short-read sequencing technology, one could use SELEX methods to characterize binding affinities of many low specificity transcription factors. (Source: BMC Biotechnology - Latest articles)

Simultaneous tracking of fly movement and gene expression using GFP

BMC Biotechnology - Latest articles — Tue, 16 Dec 2008 05:00:00 +0100

Conclusion: These experiments demonstrate that GFP can readily be used to assay longitudinally fly movement and tissue-specific patterns of gene expression. The hsp22-GFP and hsp70-GFP expression patterns were found to reflect accurately the endogenous gene expression patterns, including induction during aging and circadian periodicity. The combination of these new tracking methods with the hsp-GFP reporters revealed additional information, including a spike in hsp22 and hsp70 reporter expression preceding death, and an intriguing fly-to-fly variability in the phase of hsp70 and hsp22 reporter expression patterns. These methods allow specific temporal patterns of gene expression to be correlated with temporal patterns of animal activity, behavior and mortality. (Source: BMC Biotechnology -...

An efficient one-step site-directed deletion, insertion, single and multiple-site plasmid mutagenesis protocol

BMC Biotechnology - Latest articles — Thu, 04 Dec 2008 05:00:00 +0100

Conclusion: Our modified protocol significantly increased the efficiency of single mutation and also allowed facile large single insertions, deletions/truncations and multiple mutations in a single experiment, an option incompatible with the standard QuikChangeTM. Furthermore the new protocol required significantly less parental DNA which facilitated the DpnI digestion after the PCR amplification and enhanced the overall efficiency and reliability. Using our protocol, we generated single site, multiple single-site mutations and a combined insertion/deletion mutations. The results demonstrated that this new protocol imposed no additional reagent costs (beyond basic QuikChangeTM) but increased the overall success rates. (Source: BMC Biotechnology - Latest articles)

Reliable microRNA profiling in routinely processed formalin-fixed paraffin-embedded breast cancer specimens using fluorescence labelled bead technology

BMC Biotechnology - Latest articles — Thu, 27 Nov 2008 05:00:00 +0100

Conclusions: Comprehensive microRNA expression patterns can be reliably derived from routinely processed FFPE breast cancer specimens using fluorescence labelled bead technology. (Source: BMC Biotechnology - Latest articles)

Preparation of alginate coated chitosan microparticles for vaccine delivery

BMC Biotechnology - Latest articles — Wed, 19 Nov 2008 05:00:00 +0100

Conclusion: The prepared alginate coated chitosan microparticles, with mean diameter of about 1 micron, was suitable for oral mucosal vaccine. Moreover, alginate coating onto the surface of chitosan microparticles could modulate the release behavior of BSA from alginate coated chitosan microparticles and could effectively protect model protein (BSA) from degradation in acidic medium in vitro for at least 2h. In all, the prepared alginate coated chitosan microparticles might be an effective vehicle for oral administration of antigens. (Source: BMC Biotechnology - Latest articles)

Differential solubility of curcuminoids in serum and albumin solutions: implications for analytical and therapeutic applications

BMC Biotechnology - Latest articles — Thu, 06 Nov 2008 05:00:00 +0100

Conclusions: These results suggest the possibility of alternative therapeutic approaches by injection or infusion of relatively small amounts of curcuminoid-enriched serum. They also provide tools to reproducibly solubilize curcuminoids for analysis in cell culture applications. The differential solubility of curcuminoids achieved by different methods of solubilization offers convenient alternatives to assess the diverse biological effects contributed by curcumin and its derivatives. (Source: BMC Biotechnology - Latest articles)

Induction of multiple pleiotropic drug resistance genes in yeast engineered to produce an increased level of anti-malarial drug precursor, artemisinic acid

BMC Biotechnology - Latest articles — Tue, 04 Nov 2008 05:00:00 +0100

Conclusion: The data presented here suggest that the engineered yeast producing artemisinic acid suffers oxidative and drug-associated stresses. The use of plant-derived transporters and optimizing AMO activity may improve the yield of artemisinic acid production in the engineered yeast. (Source: BMC Biotechnology - Latest articles)

Pharmaceutical induction of ApoE secretion by multipotent mesenchymal stromal cells (MSCs).

BMC Biotechnology - Latest articles — Mon, 29 Sep 2008 04:00:00 +0100

Conclusions: The data demonstrated that pre-treatment and perhaps co-administration of MSCs homozygous for ApoE epsilon3 and dexamethasone may represent a novel therapy for severe instances of AD, atherosclerosis and other ApoE-related diseases. (Source: BMC Biotechnology - Latest articles)

Shewanella oneidensis: a new and efficient System for Expression and Maturation of heterologous [Fe-Fe] Hydrogenase from Chlamydomonas reinhardtii

BMC Biotechnology - Latest articles — Thu, 18 Sep 2008 04:00:00 +0100

Conclusion: In the present work a new efficient system for heterologous expression and maturation of C. reinhardtii hydrogenase has been developed. HydA1 of C. reinhardtii was purified and shown to contain 6 Fe atoms/molecule of protein, as expected. Using DMSO, TMAO or thiosulfate as substrates for anaerobic respiration during the cell growth, 0.4 – 0.5 mg l-1(OD600 = 1) of catalytically active HydA1 was obtained with hydrogen evolution rate of ~700 μmol H2 mg-1 min-1. (Source: BMC Biotechnology - Latest articles)

Inhibition of cell growth and invasion by epidermal growth factor-targeted phagemid particles carrying siRNA against focal adhesion kinase in the presence of hydroxycamptothecin

BMC Biotechnology - Latest articles — Thu, 18 Sep 2008 04:00:00 +0100

Conclusions: These results indicated that EGF-targeted phagemid particles might be a promising tool for anti-cancer siRNA delivery in the presence of HCPT. (Source: BMC Biotechnology - Latest articles)

The stabilizing effects of immobilization in D-amino acid oxidase from Trigonopsis variabilis

BMC Biotechnology - Latest articles — Wed, 17 Sep 2008 04:00:00 +0100

Conclusions: Multi-point covalent attachment of TvDAO onto an isoluble porous carrier provides stabilization against the denaturing effects of high temperature and exposure to a gas-liquid interface. Improvement of binding of the FAD cofactor, probably by using methods of protein engineering, would further enhance the stability of the enzyme. (Source: BMC Biotechnology - Latest articles)

Molecular analysis of chondrocytes cultured in agarose in response to dynamic compression

BMC Biotechnology - Latest articles — Mon, 15 Sep 2008 04:00:00 +0100

Conclusions: The protocols developed here offer the possibility to perform an integrated analysis of the molecular mechanisms of mechanotransduction in chondrocytes, at the gene and protein level. (Source: BMC Biotechnology - Latest articles)

Stable high volumetric production of glycosylated human recombinant IFNalpha2b in HEK293 cells

BMC Biotechnology - Latest articles — Wed, 27 Aug 2008 04:00:00 +0100

Conclusion: These results show that the HEK293 cell line is an efficient and valuable host for the production of biologically active and glycosylated human IFNa2b. (Source: BMC Biotechnology - Latest articles)

Dual-In/Out strategy for genes integration into bacterial chromosome: a novel approach to step-by-step construction of plasmid-less marker-less recombinant E. coli strains with predesigned genome structure

BMC Biotechnology - Latest articles — Tue, 12 Aug 2008 04:00:00 +0100

Conclusions: The developed Dual-In/Out strategy is a rather straightforward, but convenient combination of previously developed recombination methods: phages site-specific and general Red/ET-mediated. This new approach allows us to detail the design of future recombinant marker-less strains, carrying, in particular, rather large artificial insertions that could be difficult to introduce by usually used PCR-based Recombineering procedure. The developed strategy is simple and could be particularly useful for construction of strains for the biotechnological industry. (Source: BMC Biotechnology - Latest articles)

Display of a Maize cDNA library on baculovirus infected insect cells

BMC Biotechnology - Latest articles — Tue, 12 Aug 2008 04:00:00 +0100

Conclusion: The maize cDNA library constructed in this study relies on the novel technology of baculovirus display and is unique in currently published cDNA libraries. Produced to demonstrate proof of principle, it opens the way for the development of a eukaryotic in vivo display tool which would be ideally suited for rapid screening of the maize proteome for binding partners, such as proteins involved in hormone regulation or defence. (Source: BMC Biotechnology - Latest articles)

Detection of N-acyl homoserine lactones using a traI-luxCDABE-based biosensor as a high-throughput screening tool

BMC Biotechnology - Latest articles — Wed, 30 Jul 2008 04:00:00 +0100

Conclusions: The co-culture assays described enable the detection of AHL production in both P. aeruginosa and B. cenocepacia and should be applicable to AHL analysis in other bacterial species. The high-throughput adaptation of the liquid co-culture assay could facilitate the screening of large libraries for the identification of mutants or compounds that block the synthesis or activity of AHLs. (Source: BMC Biotechnology - Latest articles)

A comparative study of protein synthesis in in vitro Systems: from the prokaryotic reconstituted to the eukaryotic extract-based

BMC Biotechnology - Latest articles — Tue, 29 Jul 2008 04:00:00 +0100

Conclusions: The reconstituted bacterial system is sufficient but not efficient in protein synthesis. The S30 system by comparison contains additional cellular factors capable of enhancing protein translation and folding. The eukaryotic translation machinery may have evolved from its prokaryotic counterpart in order to translate more complex templates into active proteins. The data of this study may contribute to the understanding of the evolution of protein synthesis. (Source: BMC Biotechnology - Latest articles)

Critical evaluation of HPV16 gene copy number quantification by SYBR green PCR

BMC Biotechnology - Latest articles — Thu, 24 Jul 2008 04:00:00 +0100

Conclusions: Run-to-run variation in SYBR green qPCR produces unavoidable inaccuracies that should be allowed for when quantifying HPV gene copy number. While E6 copy numbers can be considered to provide a useable indication of viral loads, the E2/E6 ratio is of limited value. Previous studies may have overestimated the frequency of mixed episomal/integrant HPV infections. (Source: BMC Biotechnology - Latest articles)

Three-dimensional culture of human meniscal cells: Extracellular matrix and proteoglycan production

BMC Biotechnology - Latest articles — Thu, 26 Jun 2008 04:00:00 +0100

ConclusionMeniscal cells attached well within the 3D microenvironment and expanded with culture time. The 3D microenvironment was permissive for production of chondroitin sulfate, types I and II collagen, and to a lesser degree keratan sulfate. This microenvironment was also permissive for growth factor responsiveness, as indicated by a significant increase in proteoglycan production when cells were exposed to TGF-β (2.48 μg/ml ± 1.00, mean ± S.D., vs control levels of 1.58 ± 0.79, p < 0.0001). Knowledge of how culture microenvironments influence meniscal cell ECM production is important; the collagen sponge culture methodology provides a useful in vitro tool for study of meniscal cell biology. (Source: BMC Biotechnology - Latest articles)

Expression of HIV-1 antigens in plants as potential subunit vaccines

BMC Biotechnology - Latest articles — Mon, 23 Jun 2008 04:00:00 +0100

Conclusion: Transient agroinfiltration was best for expression of all of the recombinant proteins tested, and p24 and p17/p24 were expressed at much higher levels than Pr55Gag. Our results highlight the usefulness of plastid signal peptides in enhancing the production of recombinant proteins meant for use as vaccines. The p17/p24 protein effectively boosted T cell and humoral responses in mice primed by the DNA vaccine pTHGagC, showing that this plant-produced protein has potential for use as a vaccine. (Source: BMC Biotechnology - Latest articles)

Human IL-12 p40 as a reporter gene for high-throughput screening of engineered mouse embryonic stem cells

BMC Biotechnology - Latest articles — Tue, 03 Jun 2008 04:00:00 +0100

Conclusion: IL-12 p40 showed several advantages as a reporter gene in terms of sensitivity and ease of the detection procedure. The IL-12 p40 assay was rapid and simple, in as much as the reporter protein secreted from the transfected cells was accurately measured by ELISA using a small aliquot of the culture medium. Remarkably, expression of Il-12 p40 does not affect the pluripotency of mouse ES cells. To our knowledge, human IL-12 p40 is the first secreted reporter protein suitable for high-throughput screening of mouse ES cells. In comparison to other secreted reporters, such as the widely used alkaline phosphatase (SEAP) reporter, the IL-12 p40 reporter system offers other real advantages. (Source: BMC Biotechnology - Latest articles)

Substantially improved pharmacokinetics of recombinant human butyrylcholinesterase by fusion to human serum albumin

BMC Biotechnology - Latest articles — Fri, 16 May 2008 04:00:00 +0100

Conclusion: Both the pharmacokinetic study and the in vitro nerve agent binding and inhibition assay suggested that a fusion protein retaining both properties of huBChE and hSA is produced in vitro and in vivo. The production of the fusion protein in the milk of transgenic goats provided further evidence that sufficient quantities of BChE/hSA can be produced to serve as a cost-effective and reliable source of BChE for prophylaxis and post-exposure treatment. (Source: BMC Biotechnology - Latest articles)

Construction of a Series of Vectors for High Throughput Cloning and Expression Screening of Membrane Proteins from Mycobacterium tuberculosis

BMC Biotechnology - Latest articles — Fri, 16 May 2008 04:00:00 +0100

Conclusion: This expression platform features high throughput cloning, high flexibility for different constructs, and high efficiency for membrane protein overexpression, and is expected to be useful in membrane protein structural and functional studies. (Source: BMC Biotechnology - Latest articles)

Hydroxycamptothecin-loaded nanoparticles enhance target drug delivery and anticancer effect

BMC Biotechnology - Latest articles — Sun, 04 May 2008 04:00:00 +0100

Conclusions: Compared to the HCPT- and control-treated groups, the HCPT-loaded nanoparticle-treated group showed a more sustained release, a longer circulation time, increased delivery to tissue, and an enhanced anticancer effect. HCPT-loaded nanoparticles appear to change the pharmacokinetic behavior of HCPT in vivo. (Source: BMC Biotechnology - Latest articles)

Flow cytometric monitoring of influenza A virus infection in MDCK cells during vaccine production

BMC Biotechnology - Latest articles — Wed, 30 Apr 2008 04:00:00 +0100

Conclusions: It is shown that flow cytometry is a sensitive and robust method for the monitoring of viral infection in fixed cells from bioreactor samples. Therefore, it is a valuable addition to other detection methods of influenza virus infection such as immunotitration and RNA hybridisation. Thousands of individual cells are measured per sample. Thus, the presented method is believed to be quite independent of the concentration of infected cells (multiplicity of infection and total cell concentration) in bioreactors. This allows to perform detailed studies on factors relevant for optimization of virus yields in cell cultures. The method could also be used for process characterisation and investigations concerning reproducibility in vaccine manufacturing. (Source: BMC Biotechnology - Lat...

PCR-based gene synthesis to produce recombinant proteins for crystallization

BMC Biotechnology - Latest articles — Tue, 29 Apr 2008 04:00:00 +0100

Conclusion: We demonstrate, by using PAZ and PolA as examples, the feasibility of integrating the gene synthesis, error correction and subcloning techniques into a non-automated gene to crystal pipeline such that genes can be designed, synthesized and implemented for recombinant expression and protein crystallization. (Source: BMC Biotechnology - Latest articles)

Differential Protein Expression Following Low Temperature Culture of Suspension CHO-K1 Cells.

BMC Biotechnology - Latest articles — Tue, 22 Apr 2008 04:00:00 +0100

Conclusion: These results indicate the extent of the temperature response in CHO-K1 cells and suggest a number of key regulatory proteins and pathways that are involved in modulating the response of cells to mild hypothermia. Regulation of these identified proteins and pathways could be useful for future approaches to engineer CHO cells for improved recombinant protein production. (Source: BMC Biotechnology - Latest articles)

In vivo site-specific biotinylation of proteins within the secretory pathway using a single vector system

BMC Biotechnology - Latest articles — Fri, 18 Apr 2008 04:00:00 +0100

Conclusions: The present study shows the universal applicability to both secretory and membrane bound proteins using a single bigenic plasmid to induce the site-specific in vivo biotinylation of target molecules tagged with a short acceptor peptide. These molecules could be easily obtained from supernatants or extracts of mammalian cells and used for a wide range of biological application. (Source: BMC Biotechnology - Latest articles)

Electrospun micro- and nanofiber tubes for functional nervous regeneration in sciatic nerve transections

BMC Biotechnology - Latest articles — Fri, 11 Apr 2008 04:00:00 +0100

Conclusions: We proved that electrospun tubes, with no additional biological coating or drug loading treatment, are promising scaffolds for functional nervous regeneration. They can be knitted in meshes and various frames depending on the cytoarchitecture of the tissue to be regenerated. The versatility of this technique gives room for further scaffold improvements, tuning their mechanical properties to provide biomimetic functionalizations. Moreover, these guidance conduits can be further filled with various fillers like collagen, fibrin or self-assembling peptide gels or loaded with neurotrophic factors and seeded with cells. Electrospun scaffolds can be synthesized in different micro-architectures to regenerate lesions in other tissues like skin and bone. (Source: BMC Biotechnology - La...

Nanopolymers improve delivery of exon skipping oligonucleotides and concomitant dystrophin expression in skeletal muscle of mdx mice

BMC Biotechnology - Latest articles — Wed, 02 Apr 2008 04:00:00 +0100

Conclusion: We conclude that PEGylated PEI2K copolymers are efficient carriers for local delivery of 2-O-Me ESOs and warrant further development as potential therapeutics for treatment of DMD. (Source: BMC Biotechnology - Latest articles)

Improved production of human type II procollagen in the yeast Pichia pastoris in shake flasks by a wireless-controlled fed-batch system

BMC Biotechnology - Latest articles — Thu, 27 Mar 2008 04:00:00 +0100

Conclusions: The recommended standard protocol for methanol addition in shake flasks using pulse feeding is non-optimal and leads to repeated long phases of methanol starvation. The problem can be solved by applying the fed-batch technology. The presented wireless feeding unit, together with an on-line monitoring system offers a flexible, simple, and low-cost solution for initial optimization of the production in shake flasks which can be performed in parallel. By this way the fed-batch strategy can be applied from the early screening steps also in laboratories which do not have access to high-cost and complicated bioreactor systems. (Source: BMC Biotechnology - Latest articles)

RNAi-mediated gene silencing in tick synganglia: a proof of concept study

BMC Biotechnology - Latest articles — Wed, 26 Mar 2008 04:00:00 +0100

Conclusion: Abdominal injection of dsRNA into unfed adult female ticks appears to silence target gene expression even in the tick synganglia. The ability of dsRNA to cross the blood-brain barrier in ticks suggests that RNAi should prove to be a useful method for dissecting function of synganglia genes expressing specific neuropeptides in order to better assess their role in tick biology. (Source: BMC Biotechnology - Latest articles)

Gene targeting in adult rhesus macaque fibroblasts

BMC Biotechnology - Latest articles — Wed, 26 Mar 2008 04:00:00 +0100

Conclusions: The cell lines reported here could be used for the production of null mutant rhesus macaque models of human genetic disease using SCNT technology. In addition, given the close evolutionary relationship and biological similarity between rhesus macaques and humans, the protocols described here may prove useful in the genetic engineering of human somatic cells. (Source: BMC Biotechnology - Latest articles)

Design of a zinc finger protein binding a sequence upstream of the A20 gene

BMC Biotechnology - Latest articles — Wed, 19 Mar 2008 04:00:00 +0100

Conclusions: It is feasible to design specific zinc finger proteins by bioinformatics methods. (Source: BMC Biotechnology - Latest articles)

Production of functionally active Penicillium chrysogenum isopenicillin N synthase in the yeast Hansenula polymorpha

BMC Biotechnology - Latest articles — Wed, 19 Mar 2008 04:00:00 +0100

Conclusions: In P. chrysogenum the enzymes involved in penicillin production are compartmentalized in the cytosol and in microbodies. In this study, we focus on the cytosolic enzyme IPNS. Our data show that high amounts of functionally active IPNS enzyme can be produced in the heterologous host during cultivation at 25 degrees C, the optimal growth temperature for P. chrysogenum. This is a new step forward in the metabolic reprogramming of H. polymorpha to produce penicillin. (Source: BMC Biotechnology - Latest articles)

Comparison of different real-time PCR chemistries and their suitability for detection and quantification of genetically modified organisms

BMC Biotechnology - Latest articles — Thu, 06 Mar 2008 05:00:00 +0100

Conclusions: Although none of the chemistries significantly outperformed the others, there are certain characteristics that suggest that LNA(R) technology is an alternative to TaqMan(R) when designing assays for quantitative analysis. Because LNA(R) probes are much shorter they might be especially appropriate when high specificity is required and where the design of a common TaqMan(R) probe is difficult or even impossible due to sequence characteristics. PlexorTM on the other hand might be a method of choice for qualitative analysis when sensitivity, low cost and simplicity of use prevail. (Source: BMC Biotechnology - Latest articles)

Glycosyltransferase efficiently controls phenylpropanoid pathway

BMC Biotechnology - Latest articles — Wed, 05 Mar 2008 05:00:00 +0100

Conclusions: In plants over-expressing both the transgene for DFR and the transgene for UGT, the synthesis of phenolic acids was diverted away from the anthocyanin branch. This represents a novel approach to manipulating phenolic acids synthesis in plants. (Source: BMC Biotechnology - Latest articles)

Effects of size and topology of DNA molecules on intracellular delivery with non-viral gene carriers

BMC Biotechnology - Latest articles — Fri, 29 Feb 2008 05:00:00 +0100

Conclusions: We conclude that the nature of gene carriers was the primary determinant of cellular delivery of DNA molecules, and circular form of the DNA was more effectively processed for transgene expression. (Source: BMC Biotechnology - Latest articles)

Intracellular delivery of peptides via association with ubiquitin or SUMO-1 coupled to protein transduction domains

BMC Biotechnology - Latest articles — Fri, 29 Feb 2008 05:00:00 +0100

Conclusions : Our observations indicate that fusion of SUMO-1 to a peptide-PTD module allows generation of a stable hybrid protein that is easily produced in bacteria and which efficiently enters into cells but this property necessitates mutation of the diglycine motif at the end of SUMO-1, thereby impairing delivery of the peptide alone. (Source: BMC Biotechnology - Latest articles)

Protein kinase substrate identification on functional protein arrays

BMC Biotechnology - Latest articles — Thu, 28 Feb 2008 05:00:00 +0100

Conclusions: Functional protein microarrays are an important new tool that enables multiplex analysis of protein phosphorylation, and thus can be utilized to identify novel kinase substrates.. Integrating this technology with a systems biology approach to cell signalling will help uncover new layers in our understanding of this essential class of enzymes. (Source: BMC Biotechnology - Latest articles)

Systolic and diastolic ventricular function in zebrafish embryos: influence of norepenephrine, MS-222 and temperature

BMC Biotechnology - Latest articles — Wed, 27 Feb 2008 05:00:00 +0100

In this study, a video edge detection system was used to characterise, continuously, cardiac ventricle function in 2-5 days old zebrafish embryos embedded in 0.6% agar and examined under light microscopy at room temperature (22 degrees centigrade). Using video edge detection software (IonOptix Inc), the motion of a small region of the cardiac ventricle wall was converted to a continuous chart trace allowing analysis of wall motion amplitude (WMA) and myocardial wall velocity during systole (MWVs) and diastole (MWVd). Results: Cardiac wall motion characteristics changed progressively from day 2 to 5 (WMA, 2-days, 17.6+/-4.4 vs 5-days, 24.6+/-4.7 micrometers , p (Source: BMC Biotechnology - Latest articles)

Effect of ionic strength and presence of serum on lipoplexes structure monitorized by FRET

BMC Biotechnology - Latest articles — Tue, 26 Feb 2008 05:00:00 +0100

Conclusions: The FRET methodology described in this work allowed us to monitor stability and characterize quantitatively the structural changes (variations in interchromophore spacing distances and complexation efficiencies) undergone by DOTAP/DNA complexes in high ionic strength solutions and in presence of serum, as well as to determine the minimum amount of potentially cytotoxic cationic lipid necessary for complete coverage of DNA. This constitutes essential information regarding the thoughtful design of future in vivo applications. (Source: BMC Biotechnology - Latest articles)

Quantification of Cre-mediated recombination by a novel strategy reveals a stable extra-chromosomal deletion-circle in mice

BMC Biotechnology - Latest articles — Mon, 25 Feb 2008 05:00:00 +0100

Conclusions: eMLPA is a novel strategy which easily can be applied to measure the Cre-mediated recombination efficiency in each experimental case with high accuracy. In addition the fate of the deletion-circle can be followed simultaneously. (Source: BMC Biotechnology - Latest articles)

Enhanced gene expression from retroviral vectors

BMC Biotechnology - Latest articles — Mon, 25 Feb 2008 05:00:00 +0100

Conclusions: Retroviral vectors carrying an optimized high-level expression cassette do not produce infectious virions when introduced into packaging cells by transfection of DNA. Infectious retrovirus carrying the same cassette is readily produced when packaging cells are transfected with in vitro transcribed retroviral genomic RNA. The applications of this technique are not limited to producing the higher levels of transgene expression demonstrated here. For example, novel reporters with alternatively spliced exon-intron configurations could readily be transduced into virtually any cell. Furthermore, because the in vitro transcripts are not translated within the packaging cells, retroviruses carrying genes lethal to the packaging cells can also be produced. (Source: BMC Biotechnology - L...

Amplification of multiple genomic loci from single cells isolated by laser micro-dissection of tissues

BMC Biotechnology - Latest articles — Wed, 20 Feb 2008 05:00:00 +0100

Conclusions: Comprehensive genomic analysis of single cells from stained tissue sections opens new research opportunities for cell lineage and depth analyses, genome-wide mutation surveys, and other single cell assays. (Source: BMC Biotechnology - Latest articles)

High-throughput avian molecular sexing by SYBR green-based real-time PCR combined with melting curve analysis

BMC Biotechnology - Latest articles — Tue, 12 Feb 2008 05:00:00 +0100

Conclusions: In this study, we introduced a high-throughput avian molecular sexing technique and successfully applied it to two species. This new method holds a great potential for use in high throughput sexing of other avian species, as well. (Source: BMC Biotechnology - Latest articles)

Efficient and reproducible generation of high-expressing, stable human cell lines without need for antibiotic selection

BMC Biotechnology - Latest articles — Tue, 12 Feb 2008 05:00:00 +0100

Conclusions: We describe an improved method for developing high protein expressing stable human cell lines. These cell lines are of non-tumor origin, they are immortalized by a function not oncogenic in human and they are from an ethically accepted and easily accessible cell source. Since the cell can be easily adapted to growth in serum-free and chemically defined medium they fulfill the requirements of biopharmaceutical production processes. (Source: BMC Biotechnology - Latest articles)

Improved RNA quality and TaqMan(R) Pre-amplification method (PreAmp) to enhance expression analysis from formalin fixed paraffin embedded (FFPE) materials

BMC Biotechnology - Latest articles — Wed, 06 Feb 2008 05:00:00 +0100

Conclusions: Modification to extraction protocols has facilitated procurement of RNA that may be successfully amplified using QRT-PCR. TaqMan(R) PreAmp system is a robust and practical solution to limited quantities of RNA from FFPE extracts. (Source: BMC Biotechnology - Latest articles)