期刊
NATURE COMMUNICATIONS
卷 7, 期 -, 页码 -出版社
NATURE PUBLISHING GROUP
DOI: 10.1038/ncomms13296
关键词
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资金
- Spanish Ministry of Economy and Competitiveness [BFU2012-37969, CSD2007-00015]
- Comunidad de Madrid [S2011/BMD-2361]
- Spanish Ministry of Economy and Competitiveness
Sequencing of a single-cell genome requires DNA amplification, a process prone to introducing bias and errors into the amplified genome. Here we introduce a novel multiple displacement amplification (MDA) method based on the unique DNA primase features of Thermus thermophilus (Tth) PrimPol. TthPrimPol displays a potent primase activity preferring dNTPs as substrates unlike conventional primases. A combination of TthPrimPol's unique ability to synthesize DNA primers with the highly processive Phi29 DNA polymerase (Phi 29DNApol) enables near-complete whole genome amplification from single cells. This novel method demonstrates superior breadth and evenness of genome coverage, high reproducibility, excellent single-nucleotide variant (SNV) detection rates with low allelic dropout (ADO) and low chimera formation as exemplified by sequencing HEK293 cells. Moreover, copy number variant (CNV) calling yields superior results compared with random primer-based MDA methods. The advantages of this method, which we named TruePrime, promise to facilitate and improve single-cell genomic analysis.
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