Findings on Applied microbiology and biotechnology Reported by Investigators at University of Hong Kong (Improvement of DGGE analysis by modifications of PCR protocols for analysis of microbial community members with low abundance)
By a News Reporter-Staff News Editor at Biotech Week -- Fresh data on Biotechnology are presented in a new report. According to news reporting originating from Hong Kong, People's Republic of China, by NewsRx correspondents, research stated, "Denaturing gradient gel electrophoresis (DGGE) is a powerful technique to reveal the community structures and composition of microorganisms in complex natural environments and samples. However, positive and reproducible polymerase chain reaction (PCR) products, which are difficult to acquire for some specific samples due to low abundance of the target microorganisms, significantly impair the effective applications of DGGE."
Our news editors obtained a quote from the research from the University of Hong Kong, "Thus, nested PCR is often introduced to generate positive PCR products from the complex samples, but one problem is also introduced: The total number of thermocycling in nested PCR is usually unacceptably high, which results in skewed community structures by generation of random or mismatched PCR products on the DGGE gel, and this was demonstrated in this study. Furthermore, nested PCR could not resolve the uneven representative issue with PCR products of complex samples with unequal richness of microbial population. In order to solve the two problems in nested PCR, the general protocol was modified and improved in this study. Firstly, a general PCR procedure was used to amplify the target genes with the PCR primers without any guanine cytosine (GC) clamp, and then, the resultant PCR products were purified and diluted to 0.01 mu g ml(-1). Subsequently, the diluted PCR products were utilized as templates to amplify again with the same PCR primers with the GC clamp for 17 cycles, and the products were finally subjected to DGGE analysis. We demonstrated that this is a much more reliable approach to obtain a high quality DGGE profile with high reproducibility."
According to the news editors, the research concluded: "Thus, we recommend the adoption of this improved protocol in analyzing microorganisms of low abundance in complex samples when applying the DGGE fingerprinting technique to avoid biased results."
For more information on this research see: Improvement of DGGE analysis by modifications of PCR protocols for analysis of microbial community members with low abundance. Applied Microbiology and Biotechnology, 2014;98(12):5655-5663. Applied Microbiology and Biotechnology can be contacted at: Springer, 233 Spring St, New York, NY 10013, USA. (Springer - www.springer.com; Applied Microbiology and Biotechnology - www.springerlink.com/content/0175-7598/)
The news editors report that additional information may be obtained by contacting Y.F. Wang, University of Hong Kong, Sch Biol Sci, Lab Environm Microbiol & Toxicol, Hong Kong, Hong Kong, People's Republic of China. Additional authors for this research include F.Q. Zhang and J.D. Gu (see also Biotechnology).
Keywords for this news article include: Asia, Biotechnology, Hong Kong, People's Republic of China
Our reports deliver fact-based news of research and discoveries from around the world. Copyright 2014, NewsRx LLC