摘要
旨在基于高通量SNP芯片建立一种可评估牛早期胚胎染色体质量和生产性能的方法,为胚胎牛育种提供技术支撑。本研究通过胚胎切割技术分别对荷斯坦奶牛体内囊胚(n=27)、体外囊胚(n=21)、体外2细胞胚胎(n=6)和8细胞胚胎(n=5)进行切割取样并统计发育率,其中体内囊胚组分为1/2体内胚组(切取半个胚胎)和体内滋养层(trophoblast cells,TE)组(切取体内胚胎少量TE),体外囊胚组、体外2和8细胞胚胎分别为体外TE组(切取体外胚胎少量TE)、体外2细胞(切取体外2细胞胚胎的1个卵裂球)和8细胞组(切取体外8细胞胚胎的1个卵裂球)。切割取样后进行全基因组扩增(whole-genome amplification,WGA),对扩增成功且DNA量大于1000 ng的样品进行SNP芯片检测,芯片检出率大于90%的进行生产性能评估,低于90%的进行染色体片段缺失分析和数据填充。结果显示:1)切割部分TE后,体内TE组剩余部分和体外TE组剩余部分继续培养24 h后的发育率分别为(94.4±5.6)%和(90.5±6.6)%,而1/2体内胚组剩余部分的发育率显著降低,仅为(22.2±14.7)%。2)1/2体内胚组和体外2细胞组扩增成功率为100%,而体内TE组、体外TE组和体外8细胞组扩增的成功率分别为(94.4±5.6)%、(76.2±9.5)%和(60.0±24.5)%;与1/2体内胚组相比,体内TE组和体外TE组经WGA后DNA量均显著下降(P<0.05),且体外TE组扩增后DNA量显著低于体内TE组(P<0.05)。3)相较于1/2体内胚组(91.2±1.6)%,体内TE组(76.7±15.2)%、体外TE组(74.3±9.6)%、体外2细胞组(76.1±6.9)%、体外8细胞组(61.2±19.0)%芯片检出率均显著下降(P<0.05),且检出率均低于90%。4)以至少连续7个SNPs缺失作为染色体片段缺失筛选标准,体外TE组和体内TE组分别筛选到188个和388个染色体缺失片段,相应的缺失片段各包括46和48个基因;GO和KEGG富集分析结果显示,体外TE组缺失基因主要与细胞骨架和细胞分化相关,而体内TE组缺失基因则主要与细胞物质分泌�
This study aimed to establish a method that can assess the chromosomal quality and production performance of early bovine embryos,and provide technical support for the industrial application of bovine in vitro embryos.In this study,27 in vivo blastocyst,21 in vitro blastocyst,6 in vitro 2-cell embryos and 5 in vitro 8-cell embryos were selected,and the embryonic cell samples with different cell numbers of in vivo embryos and in vitro embryos were obtained by embryo splitting technique.Among that,in vivo embryos were divided into two groups,one was in vivo trophectoderm cells(tec) group(splitting some of the tec from the in vivo blastocyst),the other was the half in vivo embryo group(splitting half of the embryo from the in vivo blastocyst).The remaining groups were in vitro tec group(splitting some of the tec from the in vitro blastocysts),in vitro 2-cell embryo group(splitting one blastomere from the in vitro 2-cell embryo),and in vitro 8-cell embryo group(splitting one blastomere from the in vitro 8-cell embryo).After embryo splitting,whole-genome amplification(WGA) was conducted,and the samples with successful amplification(DNA amounts greater than 1 000 ng) were subjected to SNP chip detection.Data with a call rate greater than 90% were evaluated for production performance,those below 90% were subjected to chromosomal fragment deletion analysis and data filling.The results showed that:1) After splitting part of the trophectoderm cells,the developmental rates of bovine in vivo embryos(remaining portion of in vivo blastocysts after splitting some of the tec) and bovine in vitro embryos(remaining portion of in vitro blastocysts after splitting some of the tec) were(94.4±5.6)% and(90.5±6.6)%.However,the culture development rate of half of the in vivo embryo(the remaining part of the embryo after splitting half of the embryo from the in vivo blastocyst) was significantly reduced to(22.2±14.7)%.2) The success rate of DNA amplification in the half in vivo embryo group and the in vitro 2-cell group were 100%,while
作者
胡智辉
王欢
衡诺
巩建飞
王忆
王雅春
赵善江
朱化彬
HU Zhihui;WANG Huan;HENG Nuo;GONG Jianfei;WANG Yi;WANG Yachun;ZHAO Shanjiang;ZHU Huabin(Key Laboratory of Animal Genetics,Breeding and Reproduction of Ministry of Agriculture and Rural Affairs,Institute of Animal Science,Chinese Academy of Agricultural Sciences,Beijing 100193,China;China Agricultural University,Beijing 100193,China)
出处
《畜牧兽医学报》
CAS
CSCD
北大核心
2022年第11期3866-3879,共14页
ACTA VETERINARIA ET ZOOTECHNICA SINICA
基金
国家奶牛产业技术体系(CARS-36)
宁夏回族自治区重点研发计划(2021BEF02023)
中国农业科学院科技创新工程(ASTIP-IAS06)
甘肃重点研发计划(21YF5NJ196)。
关键词
牛胚胎
胚胎切割
单细胞扩增
SNP芯片
胚胎质量
bovine embryos
embryo splitting
single-cell amplification
SNP chips
embryo quality
