BALB/cA.Cg.SHJH hr 小鼠的培育及其相关遗传学特性分析

doi:10.12300/j.issn.1674-5817.2023.055

实验动物与比较医学 ›› 2023, Vol. 43 ›› Issue (4): 363-370.DOI: 10.12300/j.issn.1674-5817.2023.055

BALB/cA.Cg.SHJH ^hr 小鼠的培育及其相关遗传学特性分析

谈小倩¹, 杨颢², 唐慧青¹, 瞿伟¹, 李亮¹, 钱珍¹, 顾坚忠¹, 徐平¹()(), 肖君华²()()

^1.上海吉辉实验动物饲养有限公司, 上海 201611
^2.东华大学生物科学与技术研究所, 上海 201620

收稿日期:2023-04-28 修回日期:2023-06-08 出版日期:2023-08-25 发布日期:2023-09-14
通讯作者: 徐平（1964—），男，研究员，研究方向为实验动物资源和低温生物学。E-mail： pingxu@jh-labanimal.com。ORCID： 0000-0001-6744-8908；
肖君华（1968—），男，教授，研究方向为小鼠遗传学。E-mail: xiaojunhua@dhu.edu.cn。ORCID:0000-0002-1977-372X
作者简介:谈小倩（1982—），女，执业兽医师，研究方向为实验动物模型和实验动物疾病。E-mail： xiaoqian_tan@163.com
基金资助:
上海市科技创新行动计划项目“自发性早衰小鼠的种群建立及相关机制研究”(21140909100)

Creation and Analysis of Related Genetic Characteristics of BALB/cA.Cg.SHJH ^hr Mice

Xiaoqian TAN¹, Hao YANG², Huiqing TANG¹, Wei QU¹, Liang LI¹, Zhen QIAN¹, Jianzhong GU¹, Ping XU¹()(), Junhua XIAO²()()

^1.Shanghai Jihui Laboratory Animal Care Co. , Ltd. , Shanghai 201611, China
^2.Institute of Biological Science and Technology, Donghua University, Shanghai 201620, China

Received:2023-04-28 Revised:2023-06-08 Published:2023-08-25 Online:2023-09-14
Contact: XU Ping (ORCID: 0000-0001-6744-8908), E-mail: pingxu@jh-labanimal.com;
XIAO Junhua (ORCID: 0000-0002-1977-372X), E-mail: xiaojunhua@dhu.edu.cn;

摘要/Abstract

摘要：

目的将自发突变培育的SHJH ^hr 小鼠的突变Hr基因导入到遗传背景清晰的BALB/cAShjh近交系小鼠，通过突变基因筛查和遗传学特性分析为后续研究Hr基因突变导致异常表型的分子机制以及该模型的推广应用提供依据。方法采用在表型监测指导下回交-互交的育种方法，将自发突变培育的SHJH ^hr 小鼠的突变基因以同源突变导入方式导入到近交系BALB/cAShjh小鼠，经10代后培育成BALB/cA.Cg.SHJH ^hr （简称C.Cg.SHJH ^hr ）小鼠。通过多重PCR建库后二代测序的方法，分析C.Cg.SHJH ^hr 小鼠中随机分布于基因组上的90个单核苷酸多态性（single nucleotide polymorphism，SNP）检测位点基因型。然后按照GB/T 14927.1—2008的方法，对C.Cg.SHJH ^hr 小鼠中14个生化位点标记基因进行检测。最后利用全基因组外显子测序技术，检测该小鼠的突变基因。结果自2018年5月至2022年3月，共进行了10代的回交-互交，完成了C.Cg.SHJH ^hr 小鼠品系的构建。在检测的90个SNP位点中，除了rs13484115、rs13484116两个位点不同外，C.Cg.SHJH ^hr 小鼠的其他位点基因型均与受体小鼠BALB/cAShjh相同；生化位点标记基因检测结果显示，C.Cg.SHJH ^hr 小鼠的14个位点全部与受体小鼠相同；全基因组外显子测序发现，该小鼠相比受体小鼠品系存在109个位点突变，包括同义突变71个，终止密码子增加1个，错义突变37个，涉及蛋白质序列改变的基因20个（包括已报告的Hr基因）。结论成功培育了突变导入系C.Cg.SHJH ^hr 小鼠，通过全基因组外显子测序分析发现了3个主要引起表型变异的Hr突变基因及关联突变基因，为该小鼠在生物医学研究中的拓展应用提供了基础数据。

关键词: BALB/cA.Cg.SHJH ^hr 小鼠, 外显子测序, 突变基因, 遗传学特性

Abstract:

Objective To introduce the Hr gene of spontaneously mutated SHJH ^hr mice into BALB/cAShjh inbred mice with clear genetic background,and provide a basis for study on the molecular mechanism of Hr gene mutation-induced abnormal phenotype and the application of this model. Methods Using a backcross-intercross breeding method guided by phenotypic monitoring, mutant genes from SHJH ^hr mice bred by spontaneous mutation were introduced into inbred BALB/cAShjh mice by homozygous mutation introgression, and the mice were bred into BALB/cA.Cg.SHJH ^hr (abbreviated as C.Cg.SHJH ^hr ) mice after 10 generations. The genotypes of 90 single nucleotide polymorphism (SNP) detection sites were analyzed in C.Cg.SHJH ^hr mice by multiplex PCR library construction followed by next generation sequencing. Then 14 biochemical locus marker genes were detected in C.Cg.SHJH ^hr mice according to the method of GB/T 14927.1-2008. Finally, whole genome exon sequencing was utilized to detect the mutated genes in this mouse. Results From May 2018 to March 2022, a total of 10 generations of backcross-intercross were conducted to complete the construction of the C.Cg.SHJH ^hr mouse line. Among the 90 SNPs loci detected, except for rs13484115 and rs13484116, all the other loci had the same genotype as the recipient mice BALB/cAShjh. The results of biochemical marker gene detection showed that all the 14 loci of the mouse were the same as those of the recipient mouse. Whole genome exon sequencing found that the mouse had 109 site mutations compared with the recipient mouse strain, including 71 synonymous mutations, 1 stopgain, 37 missense mutations, and 20 genes involved in protein sequence alterations (including the reported Hr gene). Conclusion C.Cg.SHJH ^hr mice were created. Through exon sequencing and genetic analysis, three Hr mutated genes and associated mutated genes that mainly cause phenotypic variations were identified, which provides a basis for expanding the application of C.Cg.SHJH ^hr mice in biomedical research.

Key words: BALB/cA.Cg.SHJH ^hr mice, Exon sequencing, Mutation, Genetic characteristics

中图分类号:

R-332

谈小倩, 杨颢, 唐慧青, 瞿伟, 李亮, 钱珍, 顾坚忠, 徐平, 肖君华. BALB/cA.Cg.SHJH ^hr 小鼠的培育及其相关遗传学特性分析[J]. 实验动物与比较医学, 2023, 43(4): 363-370.

Xiaoqian TAN, Hao YANG, Huiqing TANG, Wei QU, Liang LI, Zhen QIAN, Jianzhong GU, Ping XU, Junhua XIAO. Creation and Analysis of Related Genetic Characteristics of BALB/cA.Cg.SHJH ^hr Mice[J]. Laboratory Animal and Comparative Medicine, 2023, 43(4): 363-370.

图/表 6

参考文献 24

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位点Loci	C.Cg.SHJH ^hr	BALB/cA
Car2	b	b
Ce2	a	a
Es1	b	b
Es3	a	a
Es10	a	a
Gpd1	b	b
Gpi1	a	a
Hbb	d	d
Idh1	a	a
Mod1	a	a
Akp1	b	b
Pgm1	a	a
Trf	b	b
Pep3	a	a

位点Loci	C.Cg.SHJH ^hr	BALB/cA
Car2	b	b
Ce2	a	a
Es1	b	b
Es3	a	a
Es10	a	a
Gpd1	b	b
Gpi1	a	a
Hbb	d	d
Idh1	a	a
Mod1	a	a
Akp1	b	b
Pgm1	a	a
Trf	b	b
Pep3	a	a

突变位置 Mutation position	碱基变化 Base alteration	基因 Gene	涉及疾病/细胞功能 Related diseases/cell functions	下游信号通路调控 Signal pathway regulation	PolyPhen-2计算得分 PolyPhen-2 scores
70559773	C→T	Hr	全身性脱毛，伴有丘疹性病变的心房颤动	是	1.000
73523427	A→G	Nudet15	造血干细胞和祖细胞急性损失	是	0.000
74744708	T→C	esd	甲醛分解代谢	是	0.505
74811692	T→C	Lrch1	细胞对趋化因子的反应	否	0.047
74813612	T→C
75031916	G→A	5031414D18Rik	自噬体成熟、糖原、脂质代谢过程的调节	否	0.022
75032403	C→A
75036145	A→G
75125178	A→G	Lrrc63	膜蛋白	否	-
75224148	C→T	Lcp1	肌动蛋白丝束组件	否	0.166
78510551	C→G	Akap11	体型缩小，活动能力减退	否	0.997
79401743	G→A	Mtrf1	线粒体翻译终止；耳聋，痉挛性共济失调	否	0.959
79401763	G→T	Mtrf1
79406608	A→G	Mtrf1
79427505	C→G	Kbtbd	蛋白酶体介导的泛素依赖蛋白的分解代谢	是	0.006
79452831	G→C	Kbtbd	蛋白酶体介导的泛素依赖蛋白的分解代谢	是	0.006
79573278	A→G	Elf1	RNA聚合酶Ⅱ转录的正调控	是	0.000
79595387	G→A	Sugt1	参与动粒功能	是	0.000
79637996	C→G	Lect1	内皮细胞增殖负调控	否	0.005
79656659	A→G
79656692	T→C
86966512	G→A	Diaph3	遗传性听神经病变，先天性造血障碍性贫血	是	0.958
96201334	G→T	Klhl1	肌动蛋白结合蛋白	否	0.001
88469236	C→G	Pcdh20	细胞黏附	否	0.000
99046045	G→C	Mzt1	微管蛋白复合体组成	是	-
99061539	G→C	Bora	蛋白激酶活性的激活，细胞周期过程的调控，蛋白质定位的调节	是	0.995
99068023	C→T	Bora
99068337	G→C
99068407	C→G
99068529	A→G
99068542	A→G
99072637	T→A
99072704	G→A
99084109	C→A	Dis3	CUT分解代谢，RNA加工，rRNA分解代谢	是	0.000
99099549	G→T	Dis3	CUT分解代谢，RNA加工，rRNA分解代谢	是	0.000
99101172	A→G	Pibf1	有丝分裂核分裂，细胞因子产生，蛋白质磷酸化	否	0.982

突变位置 Mutation position	碱基变化 Base alteration	基因 Gene	涉及疾病/细胞功能 Related diseases/cell functions	下游信号通路调控 Signal pathway regulation	PolyPhen-2计算得分 PolyPhen-2 scores
70559773	C→T	Hr	全身性脱毛，伴有丘疹性病变的心房颤动	是	1.000
73523427	A→G	Nudet15	造血干细胞和祖细胞急性损失	是	0.000
74744708	T→C	esd	甲醛分解代谢	是	0.505
74811692	T→C	Lrch1	细胞对趋化因子的反应	否	0.047
74813612	T→C
75031916	G→A	5031414D18Rik	自噬体成熟、糖原、脂质代谢过程的调节	否	0.022
75032403	C→A
75036145	A→G
75125178	A→G	Lrrc63	膜蛋白	否	-
75224148	C→T	Lcp1	肌动蛋白丝束组件	否	0.166
78510551	C→G	Akap11	体型缩小，活动能力减退	否	0.997
79401743	G→A	Mtrf1	线粒体翻译终止；耳聋，痉挛性共济失调	否	0.959
79401763	G→T	Mtrf1
79406608	A→G	Mtrf1
79427505	C→G	Kbtbd	蛋白酶体介导的泛素依赖蛋白的分解代谢	是	0.006
79452831	G→C	Kbtbd	蛋白酶体介导的泛素依赖蛋白的分解代谢	是	0.006
79573278	A→G	Elf1	RNA聚合酶Ⅱ转录的正调控	是	0.000
79595387	G→A	Sugt1	参与动粒功能	是	0.000
79637996	C→G	Lect1	内皮细胞增殖负调控	否	0.005
79656659	A→G
79656692	T→C
86966512	G→A	Diaph3	遗传性听神经病变，先天性造血障碍性贫血	是	0.958
96201334	G→T	Klhl1	肌动蛋白结合蛋白	否	0.001
88469236	C→G	Pcdh20	细胞黏附	否	0.000
99046045	G→C	Mzt1	微管蛋白复合体组成	是	-
99061539	G→C	Bora	蛋白激酶活性的激活，细胞周期过程的调控，蛋白质定位的调节	是	0.995
99068023	C→T	Bora
99068337	G→C
99068407	C→G
99068529	A→G
99068542	A→G
99072637	T→A
99072704	G→A
99084109	C→A	Dis3	CUT分解代谢，RNA加工，rRNA分解代谢	是	0.000
99099549	G→T	Dis3	CUT分解代谢，RNA加工，rRNA分解代谢	是	0.000
99101172	A→G	Pibf1	有丝分裂核分裂，细胞因子产生，蛋白质磷酸化	否	0.982

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BALB/cA.Cg.SHJH ^hr 小鼠的培育及其相关遗传学特性分析

Creation and Analysis of Related Genetic Characteristics of BALB/cA.Cg.SHJH ^hr Mice

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BALB/cA.Cg.SHJH hr 小鼠的培育及其相关遗传学特性分析

Creation and Analysis of Related Genetic Characteristics of BALB/cA.Cg.SHJH hr Mice

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BALB/cA.Cg.SHJH ^hr 小鼠的培育及其相关遗传学特性分析

Creation and Analysis of Related Genetic Characteristics of BALB/cA.Cg.SHJH ^hr Mice