Establishment of quantitative reverse transcription-polymerase chain reaction assays for human telomerase-associated genes
Establishment of quantitative reverse transcription-polymerase chain reaction assays for human telomerase-associated genes
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DOI：
10.1016/s0009-8981(99)00188-6
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发表时间：
2000-01-05
影响因子：
5
通讯作者：
Watanabe, N
中科院分区：
文献类型：
--
作者：
Yajima, T;Yagihashi, A;Watanabe, N
端粒酶是一种合成并将（TTAGGG）（n）的重复端粒序列添加到染色体末端的酶。最近，一些端粒酶相关基因已被克隆，使研究这些基因的表达成为可能。这些基因的表达和端粒酶活性的定量比较可能有助于阐明端粒酶活性的调节。因此，我们建立了一个有效的定量逆转录聚合酶链反应（RT-PCR）检测端粒酶催化亚单位（hTERT）mRNA和端粒酶相关蛋白（TEP 1）mRNA使用TaqMan（TM）荧光检测系统。使用这种方法，我们定量hTERTmRNA和TEP 1 mRNA在两种人胰腺癌细胞系AsPC-1和PANC-1中的表达。结果表明，AsPC-1细胞hTERTmRNA和TEP 1 mRNA的表达水平分别是PANC-1细胞的1.50倍和2.31倍。这种TaqMan（TM）RT-PCR分析似乎可用于确定临床标本中hTERT和TEP 1 mRNA的量。两者合计，我们的研究结果表明，它是可以测量的主要端粒酶基因的表达：亚基。此外，可以应用该技术来确定其他类型的mRNA的量。(C)2000 Elsevier Science B. V.保留所有权利。
Telomerase is an enzyme that synthesizes and adds repetitive telomeric sequences of (TTAGGG)(n) to the ends of chromosomes. Recently, several telomerase-associated genes have been cloned, making it possible to study the expression of these genes. Quantitative comparisons of the expression of these genes and of telomerase activity might help clarify the regulation of telomerase activity. Therefore, we established the validity of a quantitative reverse transcription-polymerase chain reaction (RT-PCR) assay for the human telomerase catalytic subunit (hTERT) mRNA and telomerase associated protein (TEP1) mRNA using the TaqMan(TM) fluorogenic detection system. Using this assay, we quantitated hTERT mRNA and TEP1 mRNA expression in two human pancreatic cancer cell lines, AsPC-1 and PANC-1. Our results indicated that the levels of hTERT mRNA and TEP1 mRNA expression in AsPC-1 were 1.50 and 2.31 times higher than in PANC-1 cells. This TaqMan(TM) RT-PCR assay appears to be useful in determining the quantities of hTERT and TEP1 mRNAs in clinical specimens. Taken together, our results indicate that it is possible to measure the expression of the major telomerase genes:subunits. Furthermore it is possible to apply this technique to determine the amount of other types of mRNA. (C) 2000 Elsevier Science B.V. All rights reserved.