Optimization of DNA Base Editing Technology and Its Application in Pig Genetic Modification

doi:10.13560/j.cnki.biotech.bull.1985.2023-0445

Abstract

Abstract:

Base editing technology originated from CRISPR/Cas system is the latest site-specific gene modification technology at present. According to the function characteristics of the base editor, the base editors can be divided into cytosine base editor（CBE）, adenine base editor（ABE）, glycosylase base editor（GBE）, adenine translation base editor（AYBE）, dual base editor（DBE）and prime editor（PE）. Since the emerging of base editing technology, it has been widely used in the research of animals and plants. And it has been proven to have great application value in animals and plants for genetic improvement or disease treatment. As an important agricultural economic animal and excellent disease model, genetic improvement to pig has become very important. Base editing technology has been rapidly applied in genetic improvement of animals and plants due to its convenient operation, high efficiency, fewer by-products, and high cost-effectiveness, and even it have a high prospect of providing important support for human gene therapy. In this paper, we focus on the development, optimization, application characteristics, existing issues and future prospects of base editing technology, and briefly summarize its application in pigs. It is expected to provide reference for relevant scientific workers to understand the base editing technology.

Key words: gene editing, single base editing, pig, genetic improvement, disease model

YANG Shuai-peng, QU Zi-xiao, ZHU Xiang-xing, TANG Dong-sheng. Optimization of DNA Base Editing Technology and Its Application in Pig Genetic Modification[J]. Biotechnology Bulletin, 2024, 40(1): 127-144.

Figures/Tables 3

References 108

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名称 Name	Cas蛋白 Cas protein	工具酶 Tool enzyme	PAM序列 PAM sequence	NLS	附加原件 Additional original	密码子优化 Codon optimiz-ation	编辑窗口 Editing window
BE1^[17]	dCas9	rAPOBEC1	NGG	1×	-	-	4-8
BE2^[17]	dCas9	rAPOBEC1	NGG	1×	1×UGI	-	4-8
BE3^[17]	nCas9	rAPOBEC1	NGG	1×	1×UGI	-	4-8
BE3-R33A^[28]	nCas9	rAPOBEC1R33A	NGG	1×	1×UGI	-	5-7
BE3-R33A/K34A^[28]	nCas9	rAPOBEC1R33A/K34A	NGG	1×	1×UGI	-	5-6
Target-AID^[29]	nCas9	PmCDA1	NGG	1×	1×UGI	-	2-8
TAM^[30]	dCas9	hAIDx	NGG	1×	1×UGI	-	4-10
CRISPR-X^[31]	dCas9	hAIDΔ-MS2	NGG	1×	-	-	-50-50
ZF-hAID^[32]	ZF	hAID	less	1×	1×UGI	-	-
TALE-hAID^[32]	TALE	hAID	less	1×	1×UGI	-	-
BE4^[33]	nCas9	rAPOBEC1	NGG	1×	2×UGI	-	4-8
BE4Gam^[33]	nCas9	rAPOBEC1	NGG	1×	2×UGI 1×Gam	-	4-8
eBE^[34]	nCas9	rAPOBEC1	NGG	1×	4×UGI	-	4-8
eA3A-BE3^[35]	nCas9	hAPOBEC3A	TCR	1×	1×UGI	-	7-12
BE4max^[19]	nCas9	rAPOBEC1	NGG	2×	2×UGI	-	4-8
AncBE4max^[19]	nCas9	rAPOBEC1	NGG	2×	2×UGI	+	4-8
FNLS-BE3^[36]	nCas9	rAPOBEC1	NGG	2×	1×UGI	+	3-8
hyBE4max^[37]	nCas9	rAPOBEC1	NGG	2×	2×UGI 1×Rad51DBD	-	4-12
hyA3A-BE4max^[37]	nCas9	hA3A	TCR	2×	2×UGI 1×Rad51DBD	+	3-15
YE1-BE3^[38]	nCas9	rAPOBEC1-YE1	NGG	1×	1×UGI	-	4-7
YE2-BE3^[38]	nCas9	rAPOBEC1-YE2	NGG	1×	1×UGI	-	5-6
YEE-BE3^[38]	nCas9	rAPOBEC1-YEE	NGG	1×	1×UGI	-	5-6
EE-BE3^[38]	nCas9	rAPOBEC1-EE	NGG	1×	1×UGI	-	5-5
CP-CBEmax^[39]	nCas9	rAPOBEC1	NGG	1×	2×UGI	-	4-11
SaBE3^[38]	nSaCas9	rAPOBEC1	NNGRRT	1×	1×UGI	-	3-12
VQR-BE3^[38]	VQR-Cas9nCas9	rAPOBEC1	NGA	1×	1×UGI	-	4-11
EQR-BE3^[38]	EQR-Cas9nCas9	rAPOBEC1	NGAG	1×	1×UGI	-	4-11
VRER-BE3^[38]	VRER-Cas9nCas9	rAPOBEC1	NGCG	1×	1×UGI	-	3-10
SaKKH-BE3^[38]	nSaKKHCas9	rAPOBEC1	NNNRRT	1×	1×UGI	-	3-12
XBT3^[38]	nxCas9	rAPOBEC1	NG/GAA/GAT	1×	1×UGI	-	4-8
SpCas9-NG^[40]	nSpCas9-NG	rAPOBEC1	NG	1×	1×UGI	-	4-8
dCpf1-BE^[41]	dCpf1（Cas12a）	rAPOBEC1	TTTV	3×	4×UGI	-	8-13
enCas12a-A3A-Y130F^[42]	LbCas12a	hA3A	NTTN/TYCN/TRTV	1×	1×UGI	+	7-12
RR-A3A-Y130F^[42]	LbCas12a	hA3A	NTTN/TYCN/TRTV	1×	1×UGI	+	7-12
RVR-A3A-Y130F^[42]	LbCas12a	hA3A	NTTN/TYCN/TRTV	1×	1×UGI	+	7-12
HF-BE3^[43]	HF1-nCas9	rAPOBEC1	NGG	1×	1×UGI	-	4-8
TBEs^[44]	nCas9	rAPOBEC1	NGG	1×	1×UGI 1×dCDI	-	4-8
SpG-CBE^[45]	SpG	rAPOBEC1	NGN	1×	1×UGI	-	4-8
SpRY-CBE^[45]	SpRY	rAPOBEC1	NRN	1×	1×UGI	-	4-8
ABE7.10^[18]	nCas9	TadA-TadA*	NGG	1×	-	-	4-7
ABEmax^[19]	nCas9	TadA-TadA*	NGG	2×	-	+	4-7
CP-ABEmax^[39]	CP-nCas9	TadA-TadA*	NGG	2×	-	+	4-12
ABEmaxAW^[46]	nCas9	TadA-E59A-TadA*-V106W	NGG	2×	-	+	4-8
ABEmaxQW^[46]	nCas9	TadA-E59Q-TadA*-V106W	NGG	2×	-	+	4-8
ABE7.10D53E^[47]	nCas9	TadA-D53E-TadA*-D53E	NGG	1×	-	-	4-8
ABE7.10F148A^[47]	nCas9	TadA-F148A-TadA*-F148A	NGG	1×	-	-	5
VQR-SpCas9-ABEmax^[39]	nVQR-SpCas9	TadA-TadA*	NGA	2×	-	+	4-8
VRER-SpCas9-ABEmax^[39]	nVRER-SpCas9	TadA-TadA*	NGCG	2×	-	+	4-8
VRQR-SpCas9-ABEmax^[39]	nVRQR-SpCas9	TadA-TadA*	NGA	2×	-	+	4-8
SaABEmax^[39]	nSaCas9	TadA-TadA*	NNGRRT	2×	-	+	4-14
SaKKH-ABEmax^[39]	nSaKKH	TadA-TadA*	NNNRRT	2×	-	+	4-14
ScCas9-ABE7.10^[48]	nScCas9	TadA-TadA*	NNGN	1×	-	-	4-7
xABEmax^[39]	nxCas9	TadA-TadA*	NG	2×		+	4-8
ABE-NG-S^[49]	SpCas9-NG	TadA*	NG	1×	-	+	4-7
SpG-ABEmax^[45]	SpG	TadA-TadA*	NGN	2×	-	-	4-7
SpRY-ABEmax^[45]	SpRY	TadA-TadA*	NRN	2×	-	-	4-7
ABE8e^[50]	nCas9	TadA	NGG	1×	-	-	3-11
ABE8eV106W^[50]	nCas9	TadA-V106W	NGG	1×	-	-	8-12
enCas12a-ABE8eV106W^[42]	LbCas12a	TadA-V106W	TTTV	1×	-	+	8-12
RR-ABE8eV106W^[42]	LbCas12a	TadA-V106W	TTTV	1×	-	+	8-12
RVR- ABE8eV106W^[42]	LbCas12a	TadA-V106W	TTTV	1×	-	+	8-12
TALE-ABE^[51]	TALE	TadA-F148A-TadA*-F148A	less	1×	-	-	-
TALE-CBE^[52]	nCas9	rAPOBEC1-YE1	less	1×	1×UGI	-	-
GBE^[20,53]	nCas9	rAPOBEC1	NGG	1×	1×eUNG	-	4-8
OPTI-CGBE^[54]	nCas9	rAPOBEC1	WCW	1×	1×UNG	+	4-7
eA3A-OPTI-CGBEs^[54]	nCas9	APOBEC3A	TCN	1×	1×UNG	+	4-7
hA3G-OPTI-CGBEs^[54]	nCas9	APOBEC3G	CCN	1×	1×UNG	+	4-7
hA3G-CTD-OPTI-CGBEs^[54]	nCas9	APOBEC3G	CCN	1×	1×UNG	+	4-7
AYBEv0.1^[21]	nCas9	TadA	NGG	1×	1×MPG	+	3-11
AYBEv0.2^[21]	nCas9	TadA	NGG	1×	1×MPG	+	3-11
AYBEv0.3^[21]	nCas9	TadA	NGG	1×	1×MPG	+	3-11

名称 Name	Cas蛋白 Cas protein	工具酶 Tool enzyme	PAM序列 PAM sequence	NLS	附加原件 Additional original	密码子优化 Codon optimiz-ation	编辑窗口 Editing window
BE1^[17]	dCas9	rAPOBEC1	NGG	1×	-	-	4-8
BE2^[17]	dCas9	rAPOBEC1	NGG	1×	1×UGI	-	4-8
BE3^[17]	nCas9	rAPOBEC1	NGG	1×	1×UGI	-	4-8
BE3-R33A^[28]	nCas9	rAPOBEC1R33A	NGG	1×	1×UGI	-	5-7
BE3-R33A/K34A^[28]	nCas9	rAPOBEC1R33A/K34A	NGG	1×	1×UGI	-	5-6
Target-AID^[29]	nCas9	PmCDA1	NGG	1×	1×UGI	-	2-8
TAM^[30]	dCas9	hAIDx	NGG	1×	1×UGI	-	4-10
CRISPR-X^[31]	dCas9	hAIDΔ-MS2	NGG	1×	-	-	-50-50
ZF-hAID^[32]	ZF	hAID	less	1×	1×UGI	-	-
TALE-hAID^[32]	TALE	hAID	less	1×	1×UGI	-	-
BE4^[33]	nCas9	rAPOBEC1	NGG	1×	2×UGI	-	4-8
BE4Gam^[33]	nCas9	rAPOBEC1	NGG	1×	2×UGI 1×Gam	-	4-8
eBE^[34]	nCas9	rAPOBEC1	NGG	1×	4×UGI	-	4-8
eA3A-BE3^[35]	nCas9	hAPOBEC3A	TCR	1×	1×UGI	-	7-12
BE4max^[19]	nCas9	rAPOBEC1	NGG	2×	2×UGI	-	4-8
AncBE4max^[19]	nCas9	rAPOBEC1	NGG	2×	2×UGI	+	4-8
FNLS-BE3^[36]	nCas9	rAPOBEC1	NGG	2×	1×UGI	+	3-8
hyBE4max^[37]	nCas9	rAPOBEC1	NGG	2×	2×UGI 1×Rad51DBD	-	4-12
hyA3A-BE4max^[37]	nCas9	hA3A	TCR	2×	2×UGI 1×Rad51DBD	+	3-15
YE1-BE3^[38]	nCas9	rAPOBEC1-YE1	NGG	1×	1×UGI	-	4-7
YE2-BE3^[38]	nCas9	rAPOBEC1-YE2	NGG	1×	1×UGI	-	5-6
YEE-BE3^[38]	nCas9	rAPOBEC1-YEE	NGG	1×	1×UGI	-	5-6
EE-BE3^[38]	nCas9	rAPOBEC1-EE	NGG	1×	1×UGI	-	5-5
CP-CBEmax^[39]	nCas9	rAPOBEC1	NGG	1×	2×UGI	-	4-11
SaBE3^[38]	nSaCas9	rAPOBEC1	NNGRRT	1×	1×UGI	-	3-12
VQR-BE3^[38]	VQR-Cas9nCas9	rAPOBEC1	NGA	1×	1×UGI	-	4-11
EQR-BE3^[38]	EQR-Cas9nCas9	rAPOBEC1	NGAG	1×	1×UGI	-	4-11
VRER-BE3^[38]	VRER-Cas9nCas9	rAPOBEC1	NGCG	1×	1×UGI	-	3-10
SaKKH-BE3^[38]	nSaKKHCas9	rAPOBEC1	NNNRRT	1×	1×UGI	-	3-12
XBT3^[38]	nxCas9	rAPOBEC1	NG/GAA/GAT	1×	1×UGI	-	4-8
SpCas9-NG^[40]	nSpCas9-NG	rAPOBEC1	NG	1×	1×UGI	-	4-8
dCpf1-BE^[41]	dCpf1（Cas12a）	rAPOBEC1	TTTV	3×	4×UGI	-	8-13
enCas12a-A3A-Y130F^[42]	LbCas12a	hA3A	NTTN/TYCN/TRTV	1×	1×UGI	+	7-12
RR-A3A-Y130F^[42]	LbCas12a	hA3A	NTTN/TYCN/TRTV	1×	1×UGI	+	7-12
RVR-A3A-Y130F^[42]	LbCas12a	hA3A	NTTN/TYCN/TRTV	1×	1×UGI	+	7-12
HF-BE3^[43]	HF1-nCas9	rAPOBEC1	NGG	1×	1×UGI	-	4-8
TBEs^[44]	nCas9	rAPOBEC1	NGG	1×	1×UGI 1×dCDI	-	4-8
SpG-CBE^[45]	SpG	rAPOBEC1	NGN	1×	1×UGI	-	4-8
SpRY-CBE^[45]	SpRY	rAPOBEC1	NRN	1×	1×UGI	-	4-8
ABE7.10^[18]	nCas9	TadA-TadA*	NGG	1×	-	-	4-7
ABEmax^[19]	nCas9	TadA-TadA*	NGG	2×	-	+	4-7
CP-ABEmax^[39]	CP-nCas9	TadA-TadA*	NGG	2×	-	+	4-12
ABEmaxAW^[46]	nCas9	TadA-E59A-TadA*-V106W	NGG	2×	-	+	4-8
ABEmaxQW^[46]	nCas9	TadA-E59Q-TadA*-V106W	NGG	2×	-	+	4-8
ABE7.10D53E^[47]	nCas9	TadA-D53E-TadA*-D53E	NGG	1×	-	-	4-8
ABE7.10F148A^[47]	nCas9	TadA-F148A-TadA*-F148A	NGG	1×	-	-	5
VQR-SpCas9-ABEmax^[39]	nVQR-SpCas9	TadA-TadA*	NGA	2×	-	+	4-8
VRER-SpCas9-ABEmax^[39]	nVRER-SpCas9	TadA-TadA*	NGCG	2×	-	+	4-8
VRQR-SpCas9-ABEmax^[39]	nVRQR-SpCas9	TadA-TadA*	NGA	2×	-	+	4-8
SaABEmax^[39]	nSaCas9	TadA-TadA*	NNGRRT	2×	-	+	4-14
SaKKH-ABEmax^[39]	nSaKKH	TadA-TadA*	NNNRRT	2×	-	+	4-14
ScCas9-ABE7.10^[48]	nScCas9	TadA-TadA*	NNGN	1×	-	-	4-7
xABEmax^[39]	nxCas9	TadA-TadA*	NG	2×		+	4-8
ABE-NG-S^[49]	SpCas9-NG	TadA*	NG	1×	-	+	4-7
SpG-ABEmax^[45]	SpG	TadA-TadA*	NGN	2×	-	-	4-7
SpRY-ABEmax^[45]	SpRY	TadA-TadA*	NRN	2×	-	-	4-7
ABE8e^[50]	nCas9	TadA	NGG	1×	-	-	3-11
ABE8eV106W^[50]	nCas9	TadA-V106W	NGG	1×	-	-	8-12
enCas12a-ABE8eV106W^[42]	LbCas12a	TadA-V106W	TTTV	1×	-	+	8-12
RR-ABE8eV106W^[42]	LbCas12a	TadA-V106W	TTTV	1×	-	+	8-12
RVR- ABE8eV106W^[42]	LbCas12a	TadA-V106W	TTTV	1×	-	+	8-12
TALE-ABE^[51]	TALE	TadA-F148A-TadA*-F148A	less	1×	-	-	-
TALE-CBE^[52]	nCas9	rAPOBEC1-YE1	less	1×	1×UGI	-	-
GBE^[20,53]	nCas9	rAPOBEC1	NGG	1×	1×eUNG	-	4-8
OPTI-CGBE^[54]	nCas9	rAPOBEC1	WCW	1×	1×UNG	+	4-7
eA3A-OPTI-CGBEs^[54]	nCas9	APOBEC3A	TCN	1×	1×UNG	+	4-7
hA3G-OPTI-CGBEs^[54]	nCas9	APOBEC3G	CCN	1×	1×UNG	+	4-7
hA3G-CTD-OPTI-CGBEs^[54]	nCas9	APOBEC3G	CCN	1×	1×UNG	+	4-7
AYBEv0.1^[21]	nCas9	TadA	NGG	1×	1×MPG	+	3-11
AYBEv0.2^[21]	nCas9	TadA	NGG	1×	1×MPG	+	3-11
AYBEv0.3^[21]	nCas9	TadA	NGG	1×	1×MPG	+	3-11