Skip Navigation

Nucleic Acids Research 2005 33(20):e181; doi:10.1093/nar/gni176
This Article
Right arrow Full Text Freely available
Right arrow Print PDF (372K) Freely available
Right arrow Screen PDF (381K) Freely available
Right arrow Supplementary Material
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Commercial Re-use Guidelines
for Open Access NAR Content
Google Scholar
Right arrow Articles by Suslov, O.
Right arrow Articles by Steindler, D. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Suslov, O.
Right arrow Articles by Steindler, D. A.
Related Collections
Right arrow Nucleic acid amplification
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

Published online 27 November 2005

© The Author 2005. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oxfordjournals.org

Methods Online

PCR inhibition by reverse transcriptase leads to an overestimation of amplification efficiency

Oleg Suslov and Dennis A. Steindler*

McKnight Brain Institute of the University of Florida FL, USA

*To whom correspondence should be addressed. Tel: +1 352 392 0490; Fax: +1 352 846 0185; Email: Steindler{at}mbi.ufl.edu

Received June 30, 2005. Revised September 19, 2005. Accepted October 25, 2005.

This study addresses the problem of PCR inhibition by reverse transcriptase. It has been shown that the inhibition occurs mostly when a small amount of RNA is taken for RT reaction, and it is more visible for rarely expressed transcripts. We show here that the inhibition takes place regardless of what amount of template is utilized for RT. The inhibition possesses a global nature, i.e. the amplification of any given transcript may be compromised with different levels of inhibition. The process of inhibition also explains wrongfully derived PCR amplification efficiencies, sometimes more than 100%, when the sequential dilutions of unpurified RT sample are utilized to build the calibration curve. The RT influences PCR not only by inhibiting it. When microgram(s) of RNA are taken for RT reaction, reverse transcriptase may cause overamplification of some transcripts under certain PCR conditions. The possible mechanism of RT influence on PCR is presented, and a purification method is implemented to remove the effects of RT on PCR.

Correspondence may also be addressed to Oleg Suslov. Tel: +1 352 392 6754; Fax: +1 352 392 0025; Email: osuslov{at}yahoo.com


Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 Nucleic Acids ResHome page
A. V. Lebedev, N. Paul, J. Yee, V. A. Timoshchuk, J. Shum, K. Miyagi, J. Kellum, R. I. Hogrefe, and G. Zon
Hot Start PCR with heat-activatable primers: a novel approach for improved PCR performance
Nucleic Acids Res., November 1, 2008; 36(20): e131 - e131.
[Abstract] [Full Text] [PDF]

Home page
 J EndocrinolHome page
P. D Maningat, P. Sen, A. L Sunehag, D. L Hadsell, and M. W Haymond
Regulation of gene expression in human mammary epithelium: effect of breast pumping
J. Endocrinol., December 1, 2007; 195(3): 503 - 511.
[Abstract] [Full Text] [PDF]


Disclaimer: Please note that abstracts for content published before 1996 were created through digital scanning and may therefore not exactly replicate the text of the original print issues. All efforts have been made to ensure accuracy, but the Publisher will not be held responsible for any remaining inaccuracies. If you require any further clarification, please contact our Customer Services Department.