Quantitative RT-PCR to evaluate in vivo expression of multiple transgenes using a common intron

被引：15

作者：

Fairman, J ^{[1
]}

Roche, L ^{[1
]}

Pieslak, I ^{[1
]}

Lay, M ^{[1
]}

Corson, S ^{[1
]}

Fox, E ^{[1
]}

Luong, C ^{[1
]}

Koe, G ^{[1
]}

Lemos, B ^{[1
]}

Grove, R ^{[1
]}

Fradkin, L ^{[1
]}

Vernachio, J ^{[1
]}

机构：

[1] Valentis Inc, Burlingame, CA 94101 USA

来源：

BIOTECHNIQUES | 1999年 / 27卷 / 03期

关键词：

D O I：

10.2144/99273rr04

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

An assay measuring RNA expression levels of a gene-encoded therapeutic must distinguish between endogenous mRNA and mRNA transcribed from the transgene. Specificity for the delivered transgene is especially critical when the treatment involves genes that are expressed in the target tissue. To facilitate uniform detection of transgene RNA without interference from endogenous mRNA, we have engineered expression vectors that include a 5' untranslated region (5' UTR) containing a synthetic intron (PGL3). The synthetic intron splice junction was the target sequence for a quantitative reverse transcription CRT)-PCR assay utilizing Taq-Man(R) technology. In this study, we demonstrate that a quantitative RT-PCR assay designed to recognize an engineered intron splice site in the 5' UTR of expression constructs effectively measures the expression level of in vivo-delivered gene therapeutics.

引用

页码：566 / +

页数：7

共 10 条

[1]

Ferre F, 1992, PCR Methods Appl, V2, P1

[2]

Freimark BD, 1998, J IMMUNOL, V160, P4580

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