BMPR1BFecB 同源突变的食蟹猴与恒河猴模型构建

doi:10.12300/j.issn.1674-5817.2026.122

实验动物与比较医学 ›› 2026, Vol. 46 ›› Issue (4): 467-475.DOI: 10.12300/j.issn.1674-5817.2026.122

• 人类疾病动物模型 • 下一篇

BMPR1B^FecB 同源突变的食蟹猴与恒河猴模型构建

李春杨¹, 刘星辰¹, 李玉琢¹, 陆勇¹^,², 王燕¹, 高长山¹, 聂艳红¹^,², 卞新颜¹, 胡超⁴, 朱家桥⁴, 孙强¹^,²^,³()

^1.中国科学院脑科学与智能技术卓越创新中心, 上海 200031
^2.中山大学香港高等研究院, 香港 999077
^3.中山大学中山医学院, 深圳 518107
^4.扬州大学兽医学院, 扬州 225009

收稿日期:2026-06-25 修回日期:2026-07-24 出版日期:2026-08-25 发布日期:2026-08-22
通讯作者:
孙强（1973—），男，博士，教授，博士生导师，研究方向：非人灵长类生殖生理。E-mail： qsun@sysuias.hk。ORCID： 0000-0002-3359-9465
作者简介:李春杨（1995—），女，硕士，实验师，研究方向：显微操作与胚胎发育。E-mail： lichunyang@ion.ac.cn；
刘星辰（1996—），女，博士，博士后，研究方向：非人灵长类生殖胚胎发育。E-mail： xcliu@ion.ac.cn；
李玉琢（1986—），男，硕士，高级实验师，研究方向：基因编辑模型构建。E-mail： liyuzhuo@bsbii.cn
基金资助:
国家重点研发计划-基础科研条件与重大科学仪器设备研发专项课题“猴辅助生殖与人工输精技术建立及特色育种和遗传标记研究”(2022YFF0710901)

Generation of Cynomolgus and Rhesus Monkey Models Carrying the BMPR1B^FecB Variant

LI Chunyang¹, LIU Xingchen¹, LI Yuzhuo¹, LU Yong¹^,², WANG Yan¹, GAO Changshan¹, NIE Yanhong¹^,², BIAN Xinyan¹, HU Chao⁴, ZHU Jiaqiao⁴, SUN Qiang¹^,²^,³()

^1.Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Shanghai 200031, China
^2.Sun Yat-sen University Institute of Advanced Studies Hong Kong, Hong Kong SAR 999077, China
^3.Zhongshan School of Medicine, Sun Yat-sen University, Shenzhen 518107, China
^4.College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China

Received:2026-06-25 Revised:2026-07-24 Published:2026-08-25 Online:2026-08-22
Correspondence to: SUN Qiang (ORCID: 0000-0002-3359-9465), E-mail: qsun@sysuias.hk

摘要/Abstract

摘要：

目的利用腺嘌呤单碱基编辑技术构建携带绵羊多胎基因（Booroola fecundity，FecB）同源突变的骨形态发生蛋白受体1B（bone morphogenetic protein receptor type 1B，BMPR1B）基因（即BMPR1B^FecB 同源突变）的食蟹猴和恒河猴模型，以探索提高实验猴繁殖性能的遗传改良策略。方法针对食蟹猴和恒河猴的BMPR1B基因c.746A>G位点设计4条单链向导RNA（single-guide RNAs，sgRNAs），同时结合ABE8e-SpRY单碱基编辑系统，将该编辑体系显微注射至两种猴的受精卵中。注射后的胚胎经体外培养，采用巢式PCR扩增检测目的位点碱基编辑效率，筛选高效率的sgRNA。随后将发育正常的基因编辑胚胎移植入代孕母猴输卵管内，待新生猴出生7 d后采集耳部皮肤和外周血液，进行基因型鉴定和嵌合率分析，并监测基因编辑猴的生长发育和健康状态。结果从4条候选sgRNAs中筛选到c.746A>G单碱基编辑效率达100%的sgRNA4，并通过胚胎移植获得存活仔猴17只，其中食蟹猴8只，恒河猴9只。基因型检测结果显示，8只新生食蟹猴均携带c.746A>G点突变，编辑效率为100%（8/8），其中嵌合体发生率为37.50%（3/8）；9只新生恒河猴中8只携带c.746A>G突变，编辑效率为88.89%（8/9），其中嵌合体发生率为75.00%（6/8）。结论 ABE8e-SpRY单碱基编辑系统可在食蟹猴和恒河猴胚胎中实现BMPR1B基因c.746A>G（p.Q249R）位点的定点编辑，成功建立了BMPR1B^FecB 同源突变实验猴模型，为后续在非人灵长类动物水平研究BMPR1B^FecB 同源突变对繁殖力的调控作用，以及探索实验猴繁殖性能遗传改良策略提供了重要的模型基础。

关键词: 骨形态发生蛋白受体1B, 单碱基编辑, BMPR1B^FecB 同源突变, 食蟹猴, 恒河猴

Abstract:

Objective To generate cynomolgus and rhesus monkey models carrying the Booroola fecundity (FecB) variant of bone morphogenetic protein receptor type 1B (BMPR1B) gene (designated BMPR1B^FecB ) using adenine base edit technology and to explore genetic improvement strategies for enhancing the reproductive performance of laboratory macaques. Methods Four single-guide RNAs (sgRNAs) were designed to target the BMPR1B c.746A>G locus in cynomolgus and rhesus monkeys. The sgRNAs were combined with the ABE8e-SpRY base editor, and the resulting editing complexes were microinjected into zygotes of both macaque species. Following in vitro culture of the injected embryos, nested PCR amplification was performed to detect the base-editing efficiency at the target site to screen for highly efficient sgRNAs. Subsequently, normally developing gene-edited embryos were transferred into the oviducts of surrogates. Seven days after birth, ear skin or peripheral blood samples were collected from the newborn monkeys for genotyping and mosaicism analysis, and the growth, development, and health status of the gene-edited monkeys were monitored. Results Among the four candidate sgRNAs, sgRNA4 was identified as achieving 100% editing efficiency at the c.746A>G site. A total of 17 live monkey offspring were obtained via embryo transfer, including 8 cynomolgus monkeys and 9 rhesus monkeys. Genotyping results showed that all 8 newborn cynomolgus monkeys carried the c.746A>G point mutation, with an editing efficiency of 100% (8/8), of which 37.50% (3/8) were mosaics. Among the 9 newborn rhesus monkeys, 8 individuals carried the c.746A>G mutation, with an editing efficiency of 88.89% (8/9), of which 75.00% (6/8) were mosaics. Conclusion The ABE8e-SpRY base-editing system enables precise editing at the c.746A>G (p.Q249R) locus of the BMPR1B gene in both cynomolgus and rhesus monkey embryos, resulting in the successful establishment of laboratory monkey models carrying the BMPR1B^FecB variant. These models provide a valuable foundation for future studies of the regulatory role of the BMPR1B^FecB variant in reproductive performance at the non?human primate level, as well as for exploring genetic improvement strategies to enhance reproductive performance in laboratory monkeys.

Key words: Bone morphogenetic protein receptor type 1B, Base editing, BMPR1B^FecB variant, Cynomolgus monkey, Rhesus monkey

中图分类号:

Q95-33

李春杨,刘星辰,李玉琢,等. BMPR1B^FecB 同源突变的食蟹猴与恒河猴模型构建[J]. 实验动物与比较医学, 2026, 46(4): 467-475. DOI: 10.12300/j.issn.1674-5817.2026.122.

LI Chunyang,LIU Xingchen,LI Yuzhuo,et al. Generation of Cynomolgus and Rhesus Monkey Models Carrying the BMPR1B^FecB Variant[J]. Laboratory Animal and Comparative Medicine, 2026, 46(4): 467-475. DOI: 10.12300/j.issn.1674-5817.2026.122.

图/表 6

注：A，靶位点的sgRNAs设计；B， 4条sgRNAs的编辑结果，红框内为靶位点。BMPR1B，骨形态发生蛋白受体1B；WT，野生型；Q249R，c.746A>G突变；sgRNA，单链向导RNA。
">

图 1 针对BMPR1B基因c.746A>G靶位点的sgRNA设计与筛选
注：A，靶位点的sgRNAs设计；B， 4条sgRNAs的编辑结果，红框内为靶位点。BMPR1B，骨形态发生蛋白受体1B；WT，野生型；Q249R，c.746A>G突变；sgRNA，单链向导RNA。

Figure 1 Design and screening of sgRNAs targeting the c.746A>G locus in BMPR1B gene
Note： A， Design of target-site sgRNAs； B， Editing results of the 4 sgRNAs， and the target loci are indicated within red boxes. BMPR1B， bone morphogenetic protein receptor type 1B； WT， wild type； Q249R， c.746A>G mutation； sgRNA， single-guide RNA.

表1 显微注射获得的猴胚胎中不同sgRNA的单碱基编辑效率统计

Table 1 Base-editing efficiencies of different sgRNAs in monkey embryos obtained by microinjection

sgRNA编号 sgRNA ID	编辑物种 Edited species	靶位点编辑效率/% Target-site editing efficiency/%	纯合编辑效率/% Homozygous editing efficiency/%	旁位编辑 Bystander editing
sgRNA1	MFA	28.57	0.00	Yes
sgRNA2	MFA	70.00	0.00	No
sgRNA3	MFA	0.00	0.00	Yes
sgRNA4	MFA	100.00	55.56	No
sgRNA4	MMU	100.00	91.67	No

表2 BMPR1B^FecB 基因编辑胚胎的移植和妊娠结局统计

Table 2 Embryo transfer and pregnancy outcomes for BMPR1B^FecB gene-edited embryos

物种

Species

移植胚胎数/枚

Transferred embryos

移植受体数/只

Surrogates

妊娠母体

数/只

Pregnancies

妊娠率/%

Pregnancy rate/%

胎儿数/只

Fetuses

活产数/只

Live births

活产率/%

Live birth rate/%

存活数/只

Surviving offspring

编辑数/只

Edited offspring

编辑效率/%

Editing efficiency/%

注：A，食蟹猴#0673耳尖及血液的基因型；B，恒河猴#H0806耳尖及血液的基因型。红框内为靶位点。MFA为食蟹猴，MMU为恒河猴，WT表示野生型。
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图 2 BMPR1B^FecB 基因编辑猴出生个体的基因型示例
注：A，食蟹猴#0673耳尖及血液的基因型；B，恒河猴#H0806耳尖及血液的基因型。红框内为靶位点。MFA为食蟹猴，MMU为恒河猴，WT表示野生型。

Figure 2 Representative genotypes of newborn BMPR1B^FecB gene-edited monkeys
Note： A， Genotypes of ear-tip tissue and blood samples from cynomolgus monkey #0673； B， Genotypes of ear-tip tissue and blood samples from rhesus monkey #H0806. Target loci are highlighted in red boxes. MFA， Macaca fascicularis；MMU， Macaca mulatta；WT， wild type.

表3 BMPR1B^FecB 基因编辑猴的存活个体嵌合比例

Table 3 Mosaic ratios in surviving BMPR1B^FecB gene-edited monkeys

动物编号 Animal ID	性别 Sex	基因型 Genotype	个体嵌合率/% Mosaicism rate/%
0673	Male	Hom	100.00
0677	Male	Hom	100.00
0684	Male	Hom	100.00
0687	Female	Hom	100.00
0689	Female	Mos	75.36
0693	Male	Mos	97.18
0782	Female	Mos	92.50
0783	Female	Hom	100.00
H0507	Male	Mos	62.77
H0509	Male	Mos	52.34
H0611	Male	Mos	88.51
H0813	Male	WT	0.00
H0815	Male	Mos	48.65
H0806	Female	Hom	100.00
H0819	Male	Hom	100.00
H0823	Male	Mos	67.95
H0911	Male	Mos	41.59

注：A，BMPR1B^FecB 基因编辑食蟹猴（雌性，3月龄，双胞胎）；B，BMPR1B^FecB 基因编辑恒河猴（雌性，3月龄）。
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图 3 健康存活的BMPR1B^FecB 基因编辑猴
注：A，BMPR1B^FecB 基因编辑食蟹猴（雌性，3月龄，双胞胎）；B，BMPR1B^FecB 基因编辑恒河猴（雌性，3月龄）。

Figure 3 Healthy surviving BMPR1B^FecB gene-edited monkeys
Note： A， BMPR1B^FecB gene-edited cynomolgus monkeys （females，3-month-old， twins）； B， BMPR1B^FecB gene-edited rhesus monkey （female， 3-month-old）.

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BMPR1B^FecB 同源突变的食蟹猴与恒河猴模型构建

Generation of Cynomolgus and Rhesus Monkey Models Carrying the BMPR1B^FecB Variant

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BMPR1BFecB 同源突变的食蟹猴与恒河猴模型构建

Generation of Cynomolgus and Rhesus Monkey Models Carrying the BMPR1BFecB Variant

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BMPR1B^FecB 同源突变的食蟹猴与恒河猴模型构建

Generation of Cynomolgus and Rhesus Monkey Models Carrying the BMPR1B^FecB Variant