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Limitations imposed by heteroduplex formation on quantitative RT-PCR.

Abstract：

:RT-PCR, a rapidly emerging technique for the detection of RNA, is being used by many investigators to quantify small amounts of RNA. Accurate quantification of RNA content has been facilitated by the use of competitive amplicons as internal controls. We demonstrate that losses in sensitivity and accuracy are associated with an internal standard having sequence similarity to the primary amplicon. Analysis of PCR products under non-denaturing and denaturing conditions provided evidence that these losses were associated with heteroduplex formation. Subsequent analysis of factors associated with heteroduplex formation provides insights for future development of competitive assays. Assay considerations that can minimize limitations associated with competitive PCR protocols are discussed.

journal_name

Biochem Biophys Res Commun

journal_title

Biochemical and biophysical research communications

authors

Henley WN,Schuebel KE,Nielsen DA

doi

10.1006/bbrc.1996.1319

subject

Has Abstract

pub_date

1996-09-04 00:00:00

pages

113-7

issue

1

eissn

0006-291X

issn

1090-2104

pii

S0006-291X(96)91319-4

journal_volume

226

pub_type

杂志文章

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