转录测序：一种使用RNA聚合酶进行DNA测序的方法。
Transcriptional sequencing: A method for DNA sequencing using RNA polymerase.
作者信息
Sasaki N, Izawa M, Watahiki M, Ozawa K, Tanaka T, Yoneda Y, Matsuura S, Carninci P, Muramatsu M, Okazaki Y, Hayashizaki Y
机构信息
Genome Science Laboratory, Tsukuba Life Science Center, The Institute of Physical and Chemical Research, 3-1-1 Koyadai, Tsukuba, Ibaraki 305, Japan.
出版信息
Proc Natl Acad Sci U S A. 1998 Mar 31;95(7):3455-60. doi: 10.1073/pnas.95.7.3455.
Abstract
We have developed a sequencing method based on the RNA polymerase chain termination reaction with rhodamine dye attached to 3'-deoxynucleoside triphosphate (3'-dNTP). This method enables us to conduct a rapid isothermal sequencing reaction in <30 min, to reduce the amount of template required, and to do PCR direct sequencing without the elimination of primers and 2'-dNTP, which disturbs the Sanger sequencing reaction. An accurate and longer read length was made possible by newly designed four-color dye-3'-dNTPs and mutated RNA polymerase with an improved incorporation rate of 3'-dNTP. This method should be useful for large-scale sequencing in genome projects and clinical diagnosis.
摘要
我们开发了一种基于RNA聚合酶链终止反应的测序方法，该反应使用附着有罗丹明染料的3'-脱氧核苷三磷酸（3'-dNTP）。这种方法使我们能够在<30分钟内进行快速等温测序反应，减少所需模板的量，并且无需去除会干扰桑格测序反应的引物和2'-dNTP即可进行PCR直接测序。通过新设计的四色染料-3'-dNTP和具有提高的3'-dNTP掺入率的突变RNA聚合酶，实现了准确且更长的读长。该方法应有助于基因组计划中的大规模测序和临床诊断。
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