JOURNAL ARTICLE
RESEARCH SUPPORT, U.S. GOV'T, NON-P.H.S.
Add like
Add dislike
Add to saved papers

Soil microbial community analysis using two-dimensional polyacrylamide gel electrophoresis of the bacterial ribosomal internal transcribed spacer regions.

Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) of digested genomic DNA has been previously used in comparative genomics studies of closely related bacteria species. However, a two-dimensional gel electrophoresis approach for examining microbial community structures in environmental samples has not yet been developed. We determined that it is theoretically possible to separate internal transcribed spacer regions (ITS) of bacterial communities into hundreds of operational taxonomic units (OTUs) using 2D-PAGE. Application of 2D-PAGE for separating Bacterial ITS sequences that have been PCR-amplified from replicate soil samples taken from along a Zn gradient resulted in reproducible gels containing hundreds of spots. Clear differences in spot patterns were observed between soil samples that differed in both sampling location and Zn content. The number of OTUs detected using 2D-PAGE of ITS regions was much greater than that observed using Automated Ribosomal Internal Transcribed Spacer Analysis (ARISA), Terminal Restriction Fragment Length Polymorphism (T-RFLP), or Denaturing Gradient Gel Electrophoresis (DGGE). Principal Component Analysis (PCA) of community spot patterns resulted in similar groupings of samples as those obtained using other molecular methods, however, excised spots were found to contain a far lower diversity of different sequences than excised ITS bands of the same length, as determined by RFLP analysis of excision clone libraries and subsequent sequencing of DNA eluted from excised spots. This increase in resolution makes 2D-PAGE of Bacteria ITS fragments from complex microbial communities a viable method for detecting differences between highly similar communities, as well as in streamlining follow-on sequencing efforts by reducing the level of homoplasy (co-migration of heterogeneous sequences) often seen in band-based community fingerprinting methods.

Full text links

We have located links that may give you full text access.
Can't access the paper?
Try logging in through your university/institutional subscription. For a smoother one-click institutional access experience, please use our mobile app.

For the best experience, use the Read mobile app

Mobile app image

Get seemless 1-tap access through your institution/university

For the best experience, use the Read mobile app

All material on this website is protected by copyright, Copyright © 1994-2024 by WebMD LLC.
This website also contains material copyrighted by 3rd parties.

By using this service, you agree to our terms of use and privacy policy.

Your Privacy Choices Toggle icon

You can now claim free CME credits for this literature searchClaim now

Get seemless 1-tap access through your institution/university

For the best experience, use the Read mobile app