CRISPR相关转座酶及其细菌基因组编辑应用

doi:10.13560/j.cnki.biotech.bull.1985.2022-1163

摘要/Abstract

摘要：

CRISPR-Cas能够在RNA引导下靶向DNA或RNA的特定序列，改变RNA序列即可改变靶向位点，利用这一可重编程特性已开发出了各种强大的遗传学工具。最近发现CRISPR元件在进化过程中被Tn7转座子劫持，由此衍生出的CRISPR相关转座酶（CRISPR-associated transposases, CASTs）系统具有RNA引导DNA整合的能力，被部署为靶点可重编程的基因组整合工具，在大片段和多重基因整合上具有广阔的应用前景。本文追溯了CASTs的发现历程，总结了不同类型CASTs的基因座结构特点、介导基因整合的机制模型以及其在多种革兰氏阴性细菌中的部署和应用。

Abstract:

CRISPR-Cas can target a specific sequence of DNA or RNA guided by a short RNA guide. The target sites can be changed with the RNA sequences. A variety of powerful genetic tools have been developed based on this reprogramming property. Recently, it has been found that some CRISPR elements are captured by Tn7 transposons during evolution, the derived CRISPR-associated transposases（CASTs）system has the ability of RNA-guided DNA integration. Some of them has been deployed as programmable genome integration tools, which have broad application prospects in large fragments and multiple gene integration. This review traces the discovery of CASTs, summarizes the characteristics of different types of CASTs loci, the mechanism models of gene integration, and their deployments and applications in a variety of Gram-negative bacteria.

Key words: CRISPR-associated transposases, mobile genetic element, genome editing, multiplexed gene integration

周晓杰, 杨思琪, 张译文, 徐佳琪, 杨晟. CRISPR相关转座酶及其细菌基因组编辑应用[J]. 生物技术通报, 2023, 39(4): 49-58.

ZHOU Xiao-jie, YANG Si-qi, ZHANG Yi-wen, XU Jia-qi, YANG Sheng. CRISPR-associated Transposases and Their Applications in Bacterial Genome Editing[J]. Biotechnology Bulletin, 2023, 39(4): 49-58.

图/表 5

参考文献 66

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亚型 Subtype	缩写或转座子编号 Abbreviation or transposon No.	来源 Source	底盘 Chassis	最高效率 Highest efficiency	参考文献 Reference
I-F	VchCAST	Vibrio cholerae Tn6677	Escherichia coli	~100%	[41]
			Tatumella citrea	~100%	[60]
			Klebsiella oxytoca	N.D.*	[41]
			Pseudomonas putida	N.D.*	[41]
			Klebsiella michiganensis	~0.010	[64]
			Pseudomonas simiae	~0.003	[64]
			Ralstonia sp. UNC404CL21Col	~0.001	[64]
	PtrCAST	Pseudoalteromonas translucida KMM520	Escherichia coli	~100%	[34]
	AsaCAST	Aeromonas salmonicidaS44	Escherichia coli	33.4% ^&*	[35]
	Tn7000	Vibrio cholerae 4874	Escherichia coli	~1%	[36]
	Tn7001	Photobactenium iliopiscarium NCIMB 13355	Escherichia coli	~1%	[36]
	Tn7002	Vibrio sp.F12	Escherichia coli	~0.2%	[36]
	Tn7003	Vibrnio parahaemolyticusFORC 071	Escherichia coli	~2%	[36]
	Tn7004	Vibrio sp.16	Escherichia coli	~0.05%	[36]
	Tn7005	Vibrio cholerae M1517	Escherichia coli	~45%	[36]
	Tn7006	Vibrio splendidus UCD-SED10	Escherichia coli	~0.4%	[36]
	Tn7007	Alivibrio wodanis06/09/160	Escherichia coli	~40%	[36]
	Tn7008	Alivibrio sp.1S175	Escherichia coli	~0.13%	[36]
	Tn7009	Parashewanella spongiaeHJ039	Escherichia coli	~50%	[36]
	Tn7010	Photobacterium ganghwense JCM 12487	Escherichia coli	~0.4%	[36]
	Tn7011	Pseudoalteromonas sp.P1-25	Escherichia coli	~48%	[36]
	Tn7012	Pseudoalteromonas ruthenica S3245	Escherichia coli	~5%	[36]
	Tn7013	Vibrio cholerae OYP7G04	Escherichia coli	~0.1%	[36]
	Tn7014	Vibnio diazotrophicus60.6F	Escherichia coli	~30%	[36]
	Tn7015	Shewanella sp.UCD-KL21	Escherichia coli	~6%	[36]
	Tn7016	Pseudoalteromonassp.S983	Escherichia coli	~80%	[36]
	Tn7017	Endozoicomonas ascidicolaAVMARTO5	Escherichia coli	~16%	[36]
I-B	AvCAST	Anabaena variabilis	Escherichia coli	2.50%	[37]
	PmcCAST	Peltigera membranacea cyanobiont 210A	Escherichia coli	0.85%	[37]
	RoCAST	Rippkaea orientalis	Escherichia coli	N.D.*	[38]
V-K	ShCAST	Scytonema hofmanni	Escherichia coli	80%	[27]
			Sinorhizobium meliloti	~100%	[62]
			Shewanella oneidensis MR-1	100%	[61]
			Burkholderia thailandensis	~100% **	[63]
			Pseudomonas putida	~100% **	[63]
			Agrobacterium fabrum Anabaena	40% ** N.D.*	[63] [65]
	ShoCAST	Scytonema hofmannii PCC 7110	Escherichia coli	40%	[41]
	AcCAST	Anabaena cylindrica	/	N.D.*	[27]
	ShHELIX	Scytonema hofmanni	Escherichia coli	~80%	[59]
	ShoHELIX	Scytonema hofmannii PCC 7110	Escherichia coli	~50%	[59]
	AcHELIX	Anabaena cylindrica	Escherichia coli	~90%	[59]

亚型 Subtype	缩写或转座子编号 Abbreviation or transposon No.	来源 Source	底盘 Chassis	最高效率 Highest efficiency	参考文献 Reference
I-F	VchCAST	Vibrio cholerae Tn6677	Escherichia coli	~100%	[41]
			Tatumella citrea	~100%	[60]
			Klebsiella oxytoca	N.D.*	[41]
			Pseudomonas putida	N.D.*	[41]
			Klebsiella michiganensis	~0.010	[64]
			Pseudomonas simiae	~0.003	[64]
			Ralstonia sp. UNC404CL21Col	~0.001	[64]
	PtrCAST	Pseudoalteromonas translucida KMM520	Escherichia coli	~100%	[34]
	AsaCAST	Aeromonas salmonicidaS44	Escherichia coli	33.4% ^&*	[35]
	Tn7000	Vibrio cholerae 4874	Escherichia coli	~1%	[36]
	Tn7001	Photobactenium iliopiscarium NCIMB 13355	Escherichia coli	~1%	[36]
	Tn7002	Vibrio sp.F12	Escherichia coli	~0.2%	[36]
	Tn7003	Vibrnio parahaemolyticusFORC 071	Escherichia coli	~2%	[36]
	Tn7004	Vibrio sp.16	Escherichia coli	~0.05%	[36]
	Tn7005	Vibrio cholerae M1517	Escherichia coli	~45%	[36]
	Tn7006	Vibrio splendidus UCD-SED10	Escherichia coli	~0.4%	[36]
	Tn7007	Alivibrio wodanis06/09/160	Escherichia coli	~40%	[36]
	Tn7008	Alivibrio sp.1S175	Escherichia coli	~0.13%	[36]
	Tn7009	Parashewanella spongiaeHJ039	Escherichia coli	~50%	[36]
	Tn7010	Photobacterium ganghwense JCM 12487	Escherichia coli	~0.4%	[36]
	Tn7011	Pseudoalteromonas sp.P1-25	Escherichia coli	~48%	[36]
	Tn7012	Pseudoalteromonas ruthenica S3245	Escherichia coli	~5%	[36]
	Tn7013	Vibrio cholerae OYP7G04	Escherichia coli	~0.1%	[36]
	Tn7014	Vibnio diazotrophicus60.6F	Escherichia coli	~30%	[36]
	Tn7015	Shewanella sp.UCD-KL21	Escherichia coli	~6%	[36]
	Tn7016	Pseudoalteromonassp.S983	Escherichia coli	~80%	[36]
	Tn7017	Endozoicomonas ascidicolaAVMARTO5	Escherichia coli	~16%	[36]
I-B	AvCAST	Anabaena variabilis	Escherichia coli	2.50%	[37]
	PmcCAST	Peltigera membranacea cyanobiont 210A	Escherichia coli	0.85%	[37]
	RoCAST	Rippkaea orientalis	Escherichia coli	N.D.*	[38]
V-K	ShCAST	Scytonema hofmanni	Escherichia coli	80%	[27]
			Sinorhizobium meliloti	~100%	[62]
			Shewanella oneidensis MR-1	100%	[61]
			Burkholderia thailandensis	~100% **	[63]
			Pseudomonas putida	~100% **	[63]
			Agrobacterium fabrum Anabaena	40% ** N.D.*	[63] [65]
	ShoCAST	Scytonema hofmannii PCC 7110	Escherichia coli	40%	[41]
	AcCAST	Anabaena cylindrica	/	N.D.*	[27]
	ShHELIX	Scytonema hofmanni	Escherichia coli	~80%	[59]
	ShoHELIX	Scytonema hofmannii PCC 7110	Escherichia coli	~50%	[59]
	AcHELIX	Anabaena cylindrica	Escherichia coli	~90%	[59]