Novel application of Phi29 DNA polymerase: RNA detection and analysis in vitro and in situ by target RNA-primed RCA
Novel application of Phi29 DNA polymerase: RNA detection and analysis in vitro and in situ by target RNA-primed RCA
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DOI：
10.1261/rna.1279909
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发表时间：
2009-05-01
期刊：
影响因子：
4.5
通讯作者：
Janulaitis, Arvydas
中科院分区：
文献类型：
--
作者：
Lagunavicius, Arunas;Merkiene, Egle;Janulaitis, Arvydas
我们提出了一种新的Phi 29 DNA聚合酶在基于RCA的靶RNA检测和分析中的应用。Phi 29 DNA聚合酶的3 '-> 5' RNA酶活性将靶RNA转化为引物，并且聚合酶使用该新生成的引物用于RCA起始。因此，使用靶RNA引发的RCA，锁式探针可以靶向内部RNA序列，并且可以直接分析它们的特性。我们证明了Phi 29 DNA聚合酶的核糖核酸外切活性可以成功地应用于体外和原位。这些发现扩展了检测和分析远离3 '端的RNA序列的潜力。
We present a novel Phi29 DNA polymerase application in RCA-based target RNA detection and analysis. The 3'-> 5' RNase activity of Phi29 DNA polymerase converts target RNA into a primer and the polymerase uses this newly generated primer for RCA initiation. Therefore, using target RNA-primed RCA, padlock probes may be targeted to inner RNA sequences and their peculiarities can be analyzed directly. We demonstrate that the exoribonucleolytic activity of Phi29 DNA polymerase can be successfully applied in vitro and in situ. These findings expand the potential for detection and analysis of RNA sequences distanced from 3'-end.