A tool for more specific DNA integration.
A tool for more specific DNA integration.
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用于更具体 DNA 整合的工具。
DOI：
10.1126/science.adl0863
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Sashital,DipaliG
中科院分区：
文献类型：
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作者：
Dhingra,Yukti;Sashital,DipaliG
CRISPR相关转座酶(CAST)通过将DNA结合的Cas效应蛋白与称为转座酶(，)的DNA插入酶结合起来，实现可编程的DNA整合。CAST可以为细菌和人类细胞的基因组工程引入大的DNA货物(-)。V-K型铸型由Cas效应子Cas12k、转座酶TnsB、三磷酸腺苷酶(ATPase)TnsC和接头蛋白TniQ组成。这些铸型是特别有吸引力的基因组工程工具，因为它们的大小和亚基的数量都很小，而且它们能够单向整合货物DNA。然而，V-K型CAST的一个限制是它们在全基因组范围内脱靶整合的高倾向(，，，)。在本期第784页，Georgeet al.报道称，V-K型铸型的脱靶整合独立于Cas12k发生，揭示了一种非靶向转位的机制。这种非靶向转座依赖于TnsC的浓度，这使得作者能够设计出一种特异性大大提高的CAST系统。
CRISPR-associated transposases (CASTs) enable programmable DNA integration by combining DNA-binding Cas effector proteins with DNA-inserting enzymes called transposases (, ). CASTs can introduce large DNA cargoes for genome engineering in bacterial and human cells (–). Type V-K CASTs consist of the Cas effector Cas12k, the transposase TnsB, the adenosine triphosphatase (ATPase) TnsC, and the adaptor protein TniQ. These CASTs are particularly attractive genome engineering tools owing to their minimal size and number of subunits and their ability to integrate cargo DNA unidirectionally . However, a limitation of type V-K CASTs is their high propensity for genome-wide off-target integration (, , , ). On page 784 of this issue, Georgeet al. report that off-target integration by type V-K CASTs occurs independently of Cas12k, revealing a mechanism of untargeted transposition. This untargeted transposition was dependent on TnsC concentration, which allowed the authors to engineer a CAST system with much improved specificity.