A thermostable Cas9 with increased lifetime in human plasma

被引：125

作者：

Harrington, Lucas B. ^{[1
]}

Paez-Espino, David ^{[2
]}

Staahl, Brett T. ^{[1
]}

Chen, Janice S. ^{[1
]}

Ma, Enbo ^{[1
]}

Kyrpides, Nikos C. ^{[2
]}

Doudna, Jennifer A. ^{[1
,3
,4
,5
,6
]}

机构：

[1] Univ Calif Berkeley, Dept Mol & Cell Biol, 229 Stanley Hall, Berkeley, CA 94720 USA

[2] Joint Genome Inst, Dept Energy, Walnut Creek, CA 94598 USA

[3] Univ Calif Berkeley, Dept Chem, Berkeley, CA 94720 USA

[4] Univ Calif Berkeley, Howard Hughes Med Inst, Berkeley, CA 94720 USA

[5] Univ Calif Berkeley, Innovat Genom Inst, Berkeley, CA 94720 USA

[6] Lawrence Berkeley Natl Lab, MBIB Div, Berkeley, CA 94720 USA

来源：

NATURE COMMUNICATIONS | 2017年 / 8卷

基金：

美国国家科学基金会;

关键词：

HALF-LIFE; DUAL-RNA; PROTEINS; CRISPR-CAS9; PREDICTION; EVOLUTION; GENE;

D O I：

10.1038/s41467-017-01408-4

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

CRISPR-Cas9 is a powerful technology that has enabled genome editing in a wide range of species. However, the currently developed Cas9 homologs all originate from mesophilic bacteria, making them susceptible to degradation and unsuitable for applications requiring cleavage at elevated temperatures. Here, we show that the Cas9 protein from the thermophilic bacterium Geobacillus stearothermophilus (GeoCas9) catalyzes RNA-guided DNA cleavage at elevated temperatures. GeoCas9 is active at temperatures up to 70 degrees C, compared to 45 degrees C for Streptococcus pyogenes Cas9 (SpyCas9), which expands the temperature range for CRISPR-Cas9 applications. We also found that GeoCas9 is an effective tool for editing mammalian genomes when delivered as a ribonucleoprotein (RNP) complex. Together with an increased lifetime in human plasma, the thermostable GeoCas9 provides the foundation for improved RNP delivery in vivo and expands the temperature range of CRISPR-Cas9.

引用

页数：8

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