Anchor primer associated problems in differential display reverse transcription polymerase chain reaction

Chen, Yihua and Wang, Bin and Weining, Song and Daggard, Grant (2004) Anchor primer associated problems in differential display reverse transcription polymerase chain reaction. Analytical Biochemistry, 329 (1). pp. 145-147. ISSN 0003-2697

Abstract

Differential display reverse transcription PCR (DDRT-PCR)2 was first reported by P. Liang and A.B. Paredee in 1992. The method utilized an anchor primer, including a poly(T) component with the addition of one or two select bases in reverse transcription. The first-strand cDNA was amplified via PCR using the same anchor primer and a 10-base random primer. As the DDRT-PCR method is simple and sensitive, it has become a popular technology in gene expression work. However, a major problem with the technique is its ability to produce a high rate of false-positive products. A number of approaches to overcome such problems have been suggested, including using improved gel resolution systems, performing PCR with the cDNA generated by two different reverse transcriptases, and using cytoplasmic RNA to avoid unprocessed mRNA. While there has been a focus in the literature on modification of the random primer component as a means of improving DDRT-PCR reliability, in this study we report on the impact of modifications to the traditional anchor primer component and the role of anchor primers. The addition of weight bases is important for improving the reliability of traditional anchor primer in the DDRT PCR condition and can contribute to the reproducibility of DDRT PCR products.


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Item Type: Article (Commonwealth Reporting Category C)
Refereed: Yes
Item Status: Live Archive
Additional Information: Author's version not held.
Depositing User: Ms Debbie White
Faculty / Department / School: Historic - Faculty of Sciences - Department of Biological and Physical Sciences
Date Deposited: 30 Apr 2010 12:21
Last Modified: 02 Jul 2013 23:51
Uncontrolled Keywords: DDRT-PCR, differential display reverse transcription PCR
Fields of Research (FOR2008): 10 Technology > 1001 Agricultural Biotechnology > 100199 Agricultural Biotechnology not elsewhere classified
06 Biological Sciences > 0604 Genetics > 060405 Gene Expression (incl. Microarray and other genome-wide approaches)
Socio-Economic Objective (SEO2008): E Expanding Knowledge > 97 Expanding Knowledge > 970106 Expanding Knowledge in the Biological Sciences
Identification Number or DOI: doi: 10.1016/j.ab.2004.02.030
URI: http://eprints.usq.edu.au/id/eprint/7827

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