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Taq DNA聚合酶的单个氨基酸变化可实现更快的聚合酶链式反应、逆转录和链置换。

A Single Amino Acid Change to Taq DNA Polymerase Enables Faster PCR, Reverse Transcription and Strand-Displacement.

作者信息

Barnes Wayne M, Zhang Zhian, Kermekchiev Milko B

机构信息

Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO, United States.

DNA Polymerase Technology, Inc., St. Louis, MO, United States.

出版信息

Front Bioeng Biotechnol. 2021 Jan 14;8:553474. doi: 10.3389/fbioe.2020.553474. eCollection 2020.

Abstract

A change of an aspartic acid to asparagine of Taq () DNA polymerase is a gain of function mutation that supports faster PCR: the extension times for PCR amplification can be 2-3 times shorter. Surprising results from negative controls led to the discovery of strand-displacement ability and reverse transcriptase activity of Taq D732N DNA polymerase. We demonstrate that the mutant enzyme can, by itself, catalyze RT-PCR, and RT-LAMP assays. Residue 732 is on the surface of the enzyme, not near the active site.

摘要

嗜热栖热菌(Taq)DNA聚合酶中天冬氨酸突变为天冬酰胺是一种功能获得性突变,可实现更快的聚合酶链式反应(PCR):PCR扩增的延伸时间可缩短2至3倍。阴性对照的意外结果导致发现了Taq D732N DNA聚合酶的链置换能力和逆转录酶活性。我们证明,这种突变酶自身即可催化逆转录PCR(RT-PCR)和逆转录环介导等温扩增(RT-LAMP)检测。732位残基位于酶的表面,而非活性位点附近。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/16e2/7841393/ec58687f3d94/fbioe-08-553474-g001.jpg

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