creators_name: Pandey, Janmejay
creators_name: Ganesan, K
creators_name: Jain, R K
type: article
datestamp: 2012-01-09 05:06:19
lastmod: 2012-04-03 07:12:51
metadata_visibility: show
title: Variations in T-RFLP profiles with differing chemistries of fluorescent dyes used for labeling the PCR primers.
ispublished: pub
subjects: QR
full_text_status: restricted
keywords: Microbial ecology; Community structure; Variation; T-RFLP analysis
note: Copyright of this article belongs to Elsevier Science.
abstract: Culture independent molecular methods have emerged as indispensable tools for studying microbial community structure and dynamics in natural habitats, since they allow a closer look at microbial diversity that is not reflected by culturing techniques. Terminal Restriction Fragment Length Polymorphism (T-RFLP) analysis is one of the informative and widely used techniques for such studies. However, the method has a few limitations to predict microbial community structure with significant accuracy. One of the major limitations is variation in real Terminal Restriction Fragment (TRF) length and observed TRF length. In the present study we report the generation of TRF length variations using different fluorescent dyes to label the PCR primers. T-RFLP profiles generated from primers labeled with different dyes varied significantly and led to inconsistent microbial species identification. Occurrence of such variations can have serious consequences on interpretation of the T-RFLP profiles from environmental samples representing complex microbial community. Therefore, in a T-RFLP study, the primers and labeling dye system should be carefully evaluated and optimized for an individual community under investigation. Further, it would be recommended to establish a target gene library in parallel with T-RFLP analysis to facilitate the accurate prediction of microbial community structure.
date: 2007-03
date_type: published
publication: Journal of microbiological methods
volume: 68
number: 3
publisher: Elsevier Science
pagerange: 633-8
refereed: TRUE
issn: 0167-7012
official_url: http://www.sciencedirect.com/science/article/pii/S0167701206003411
related_url_url: http://www.sciencedirect.com/science/article/pii/S0167701206003411
related_url_type: pub
citation:   Pandey, Janmejay and Ganesan, K and Jain, R K  (2007) Variations in T-RFLP profiles with differing chemistries of fluorescent dyes used for labeling the PCR primers.  Journal of microbiological methods, 68 (3).  pp. 633-8.  ISSN 0167-7012     
document_url: http://crdd.osdd.net/open/126/1/jain2007.2.pdf