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中国生物工程杂志

China Biotechnology
China Biotechnology
China Biotechnology  2023, Vol. 43 Issue (10): 32-42    DOI: 10.13523/j.cb.2304020
    
Characteristics and Application of CRISPR/Cas12a Genome Editing Technology
BAO Xin-ru1,CHEN Mao-sen1,ZHONG Jie1,QI Feng1,2,**()
1 National and Local Joint Engineering Research Center for Industrial Microbial Fermentation Technology, College of Life Science, Fujian Normal University, Fuzhou 350117, China
2 Fujian Provincial Key Laboratory of Cell Stress Response and Metabolic Regulation, Fujian Normal University, Fuzhou 350108, China
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Abstract  

The CRISPR/Cas system (clustered regularly interspaced short palindromic repeats-CRISPR associated proteins) has been developed as a new technique widely used in gene editing in recent years. It is derived from the adaptive immune system of bacteria and archaea. At present, the CRISPR system has been extensively explored, among which Class II CRISPR/Cas9 is used as an effective gene editing tool in microorganisms, plants and animal cells. Compared to typical CRISPR/Cas9, CRISPR/Cas12a, which belongs to Class II technology, has also been investigated and developed and has been receiving increasing attention due to its unique advantages. This review aims to introduce the characteristics of CRISPR/Cas12a, its differences from CRISPR/Cas9 in the biological mechanisms, as well as its application in synthetic biology and medicine so as to provide reference for the further development and application of CRISPR/Cas12a gene editing technology.

Key words： CRISPR/Cas12a      Genome editing      Transcriptional regulation     
Received: 10 April 2023      Published: 02 November 2023
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BAO Xin-ru, CHEN Mao-sen, ZHONG Jie, QI Feng. Characteristics and Application of CRISPR/Cas12a Genome Editing Technology. China Biotechnology, 2023, 43(10): 32-42.

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https://manu60.magtech.com.cn/biotech/10.13523/j.cb.2304020     OR     https://manu60.magtech.com.cn/biotech/Y2023/V43/I10/32

Fig.1 The biological mechanisms and Cas domain organizations of CRISPR/Cas (a) CRISPR/Cas9 (b) CRISPR/Cas12a
项目 CRISPR/Cas9 CRISPR/Cas12a
Cas蛋白大小 1 000~1 600 aa 1 200~1 300 aa
crRNA长度 100~200 nt 42~44 nt
crRNA类型 双链 单链
原间隔序列邻近基序 3' -GGN(SpCas9) 5' -TTN(FnCas12a)
核酸内切酶结构域 RuvC和HNH RuvC和Nuc
加工pre-crRNA RNase Ⅲ Cas12a
切口类型 平末端 黏性末端
多基因编辑效率 低 高
Table 1 The difference between CRISPR/Cas9 and CRISPR/Cas12a
Fig.2 A model for CRISPR/Cas12a-mediated transcriptional regulation
目的 工具 Cas12a蛋白来源 应用 参考文献
转录调控 dAsCas12a-SoxS(CRISPRi和CRISPRa) Acidaminococcus sp. Paenibacillus polymyx [24]
dEeCas12a(CRISPRi) Eubacterium eligens Escherichia coli MG1655
Escherichia coli DH5α		 [36]
[36]
dFn Cas12a(CRISPRi) Francisella novicida Solventogenic Clostridium [49]
dFnCas12a(CRISPRi和CRISPRa) Francisella novicida Bacillus amyloliquefaciens [50]
LbCas12a-Mxi1 (CRISPRi)
LbCas12a-VPR(CRISPRa)		 Lachnospiraceae sp.
Lachnospiraceae sp.		 Yarrowia lipolytica
Yarrowia lipolytica		 [35]
[35]
AsCas12a-VPR(CRISPRa) Acidaminococcus sp. 小鼠细胞 [51]
基因重组 FnCas12a-λ-Red Francisella novicida Escherichia coli MG1655
Halomonas bluephagenesis		 [39]
[39]
FnCas12a-ssDNA Francisella novicida Escherichia coli MG1655
Yersinia pestis KIM6+
Mycobacterium smegmatis mc2155		 [40]
[40]
[40]
SEVA-Cpf1 Francisella novicida Cyanobacteria [52]
AsCas12a-REDIT Acidaminococcus sp. 人类细胞 [41]
AsCas12a-MITI Acidaminococcus sp. 人类细胞 [42]
AsCas12a-Plus突变体
LbCas12a-Plus突变体		 Acidaminococcus sp.
Lachnospiraceae sp.		 哺乳动物细胞
哺乳动物细胞		 [53]
[53]
碱基编辑 FnCas12a-ODM Francisella novicida Corynebacterium glutamicu [54]
bsdFnCas12a-BE Francisella novicida Escherichia coli DH5a [55]
dLbCas12a-BE0 Lachnospiraceae sp. 人类细胞 [45]
dLbCas12a突变体-CEBs/ABEs Lachnospiraceae sp. 哺乳动物细胞 [46]
dLbCas12a-BEACON Lachnospiraceae sp. 哺乳动物细胞 [44]
Table 2 CRISPR/Cas12a genome editing tool
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