体内、外卵母细胞对猪体细胞克隆胚胎发育潜力的影响

doi:10.16431/j.cnki.1671-7236.2017.11.005

《中国畜牧兽医》 ›› 2017, Vol. 44 ›› Issue (11): 3143-3148.doi: 10.16431/j.cnki.1671-7236.2017.11.005

体内、外卵母细胞对猪体细胞克隆胚胎发育潜力的影响

华再东¹, 郭帅², 肖红卫¹, 任红艳¹, 张立苹¹, 葛耀文²

1. 湖北省农业科学院畜牧兽医研究所, 动物胚胎工程及分子育种湖北省重点实验室, 武汉 430064;
2. 武汉维尔鑫动物种质资源开发有限公司, 武汉 430209

修回日期:2017-07-11 出版日期:2017-11-20 发布日期:2017-11-21
通讯作者: 华再东 E-mail:zaidonghua@163.com
作者简介:华再东(1978-),男,贵州盘县人,博士,研究方向:动物繁殖生物技术
基金资助:
国家转基因生物新品种培育重大专项（2016ZX08006002-006、2016ZX08006001-005、2016ZX08010003-006）；湖北省农业科技创新中心（2016-620-000-001-027）

Effect of Oocytes in vivo and in vitro on the Developmental Potential of Porcine Somatic Cell Cloned Embryos

HUA Zai-dong¹, GUO Shuai², XIAO Hong-wei¹, REN Hong-yan¹, ZHANG Li-ping¹, GE Yao-wen²

1. Hubei Key Laboratory of Animal Embryo Engineering and Molecular Breeding, Institute of Science and Veterinary Medicine, Hubei Academy of Agriculture Sciences, Wuhan 430064, China;
2. Wuhan ViaGen Animal Breeding Resources Developing Co., Ltd., Wuhan 430209, China

Revised:2017-07-11 Online:2017-11-20 Published:2017-11-21

摘要/Abstract

摘要：

为探讨猪体内、外成熟卵母细胞对核移植重组胚胎发育能力的影响，试验通过激素促排获得体内成熟卵母细胞和收集废弃卵巢获取体外成熟的卵母细胞，分别构建核移植重组胚，比较其卵裂率、囊胚率及胚胎移植受孕情况。结果显示，PGC+PMSG+HCG组的平均排卵数（27.8枚/头）显著高于PGC+HCG （12.5枚/头）、PMSG+HCG （13.7枚/头）及自然发情组（11.5枚/头）（P<0.05），体内收集到的卵母细胞，可用于构建核移植重组胚的可用卵率均达到90%以上，与其他处理组差异不显著（P>0.05），说明通过激素处理可获得更多的可用卵母细胞，而且卵母细胞的质量没有显著差异；以体内和体外成熟卵母细胞作为核移植受体构建的克隆胚胎，二者的胚胎融合率（80.31%和79.29%）和卵裂率（90.40%和86.51%）差异均不显著（P>0.05），但来自体内成熟卵母细胞克隆的胚胎发育至囊胚期的比例显著升高（P<0.05）；将体内、外成熟卵母细胞构建的核移植重组胚分别移植代孕母猪，头平均移植30或60枚时，体内成熟卵母构建的克隆胚胎移植出生仔猪10头，而体外培养卵母细胞构建的克隆胚胎均未着床受孕，表明通过激素促排获得的卵母细胞质量更好，能显著提高克隆胚胎的囊胚率，减少胚胎移植数量，提高代孕母猪的怀孕率。

关键词: 卵母细胞; 体细胞核移植; 胚胎; 猪

Abstract:

To investigate the impact of porcine oocytes in vivo and in vitro maturation (IVM) on the development of porcine somatic cell cloned embryos,the somatic cell cloned embryos cultured in vitro and the sows were treated with hormones to collect mature oocytes in vivo,and the cleavage rate, blastocyst rate and embryo implantation were compared. The results showed that the average number of ovulation in PGC+PMSG+HCG group was significantly higher than that of PGC+HCG,PMSG+HCG and the natural estrus groups (P<0.05). The oocytes collected in vivo could be used for the construction, and the available oocytes rate reached more than 90%,and there was no significant difference among the four groups (P>0.05),which indicated that groups treated by hormone could obtain more available oocytes and the quality of oocytes was not significant different. In vivo and in vitro matured oocytes were used as nuclear transfer embryos of recombinant receptor,the fusion efficiency (80.31% and 79.29%) and cleavage rate (90.40% and 86.51%) were not significant different (P>0.05), but the proportion of in vivo matured oocytes cloned embryos developed into the blastocyst stage was significantly higher (P<0.05). The reconstructed embryos made from in vivo and in vitro matured oocytes were transplanted into surrogate sows (transferred 30 or 60 embryos),10 piglets were born in in vivo maturation of cloned embryo transfer group,while there was no implantation in in vitro maturation of cloned embryo transfer group. The results showed that high quality oocytes obtained by superovulation could significantly increase the blastocyst rate of embryos,reduce the number of embryos transferred and improve the pregnancy rate of surrogate sows.

Key words: oocytes; somatic cell nuclear transfer; embryos; sow

中图分类号:

Q813.7

华再东, 郭帅, 肖红卫, 任红艳, 张立苹, 葛耀文. 体内、外卵母细胞对猪体细胞克隆胚胎发育潜力的影响[J]. 《中国畜牧兽医》, 2017, 44(11): 3143-3148.

HUA Zai-dong, GUO Shuai, XIAO Hong-wei, REN Hong-yan, ZHANG Li-ping, GE Yao-wen. Effect of Oocytes in vivo and in vitro on the Developmental Potential of Porcine Somatic Cell Cloned Embryos[J]. , 2017, 44(11): 3143-3148.

参考文献

[1] Wilmut I,Schnieke A E,Mcwhir J,et al. Viable offspring derived from fetal and adult mammalian cells[J]. Nature,1997,385(6619):810-813.
[2] Kato Y,Tani T,Sotomaru Y,et al.Eight calves cloned from somatic cells of a single adult[J].Science,1998,282(5396):2095-2098.
[3] Cibelli J B,Stice S L,Golueke P J,et al. Cloned transgenic calves produced from nonquiescent fetal fibroblasts[J].Science,1998,280(5367):1256-1258.
[4] Baguisi A,Behboodi E,Melican D T,et al.Production of goats by somatic cell nuclear transfer[J].Nature Biotechnology,1999,17(5):456-461.
[5] Wakayama T,Perry A C,Zuccotti M,et al. Full-term development of mice from enucleted oocytes injected with cumulus cell nuclei[J].Nature,1998,394(6691):370-374.
[6] Polejaeva I A,Chen S H,Vaught T D,et al. Cloned pigs produced by nuclear transfer form adult somatic cell[J].Nature,2000,407(6800):85-90.
[7] Onishi A,Iwamoto M,Akita T,et al. Pig cloning by microinjection of fetal fibroblast nuclei[J]. Scinece,2000,289(5482):1188-1190.
[8] Huan Y J,Wu Z F,Zhang J G, et al. Alteration of the DNA methylation status of donor cells impairs the developmental competence of porcine cloned embryos[J].Journal of Reproduction & Development,2016,62(1):71-77.
[9] Dai J,Leng L Z,Lu C F,et al.Time-lapse observation and transcriptome analysis of a case with repeated multiple pronuclei after IVF/ICSI[J].J Assist Reprod Genet,2017,6(1):1-9.
[10] Scott L.The biological basis of non-invasive strategies for selection of human oocytes and embryos[J].Human Reproduction Update,2003,9(3):237-249.
[11] Niemann H, Rath D.Progress in reproductive biotechnology in swine[J].Theriogenology,2001,56(8):1291-1304.
[12] 华再东,郑新民,魏庆信,等.体细胞核移植生产巴马猪[J]. 中国畜牧兽医,2014,41(10):192-197.
[13] 郭润发.优化巴马香猪超排和利用CRISPR/Cas9技术建立PDX-1基因敲除猪的研究[D].哈尔滨:东北农业大学,2016.
[14] Lin T,Oqani R K,Lee J E,et al.Coculture with good-quality COCs enhances the maturation and development rates of poor-quality COCs[J].Theriogenology,2016,85(3):396-407.
[15] Watson A J.Oocyte cytoplasmic maturation:A key mediator of oocyte and embryo developmental competence[J].Journal of Animal Science,2007,85(13):1-3.
[16] Ata B,Shalompaz E,Chian R C,et al. In vitro maturation of oocytes as a strategy for fertility preservation[J]. Clinical Obstetrics & Gynecology,2010,53(4):775-786.
[17] Fauque P.Ovulation induction and epigenetic anomalies[J].Fertility & Sterility,2013,99(3):616-623.
[18] El H N,Haaf T. Epigenetic disturbances in in vitro cultured gametes and embryos:Implications for human assisted reproduction[J]. Fertility & Sterility,2013,99(3):632-641.
[19] Denomme M M,Mann M R. Genomic imprints as a model for the analysis of epigenetic stability during assisted reproductive technologies[J].Reproduction,2012,144(4):393-409.
[20] Piquette G N. The in vitro maturation (IVM) of human oocytes for in vitro fertilization (IVF):Is it time yet to switch to IVM-IVF?[J]. Fertility & Sterility,2006,85(4):833-835.
[21] Liu Y,Lucas-Hahn A,Petersen B,et al. Developmental competence and epigenetic profile of porcine embryos produced by two different cloning methods[J].Cell Reprogram,2017,19(3):171-179.
[22] Krisher R L, Brad A M, Herrick J R,et al. A comparative analysis of metabolism and viability in porcine oocytes during in vitro maturation[J]. Animal Reproduction Science,2007,98(1):72-96.

体内、外卵母细胞对猪体细胞克隆胚胎发育潜力的影响

Effect of Oocytes in vivo and in vitro on the Developmental Potential of Porcine Somatic Cell Cloned Embryos

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