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The study of production of chimera bovine using aggregated embryos with parthenogenetic and fertilized embryos

利用孤雌胚胎和受精胚胎聚合胚胎生产嵌合牛的研究

基本信息

批准号：
08456160
负责人：
SUZUKI Tatsuyuki
金额：
$ 4.99万
依托单位：
Yamaguchi University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1998
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08456160/
关键词：
bovine parthenogenesis embryo aggregation chimeric embryos transfer pregnant chimera clone キメラ

项目摘要

The aggregation rate of aggregated embryos that were produced in vitro without agar embedding was lower(P<0.01) than with 1.0% and 1.2% agar embedding(53-70% vs 93-95%). However, based on the number of aggragated embryos, the development rate to blastocyst was not different among the treatments. The aggregated embryos produced by injection of parthenogenetic demi-embryos to the male fertilized demi-embryos and aggregated embryos produced by aggregation of entire parthenogenetic and fertlized embryos were karyotyped. Karyotyping of aggregated embryos resulted in 53% of embryos having XX and XY chromosome plates in the same sample, 27% of embryos with XY and 20% of embryos with XX chromosome plates. The mean total cell number per blastocyst of aggregated embryos produced by aggregated with the demi-embryos was lower(P<0.01) than those produced by aggregated with entire embryos(158*65 vs 202*32).Six out of eleven recipients were diagnosed pregnant after transfer of the aggregated embryos with Holstein(fertilized with Japanese Black) and Japanese Brown(parthnogenetic embryo) : Two out of six were aborted and obtained 7 calves(3 twin). The twin males (still birth) were delivered at day 234 of gestation from recipient in which XX and XY chromosome plates were detected in each calf. The single male calf which XX and XY chromosome plates were detected was delivered at day 261 of gestation. Skin color was also used as evidence of chimerism. The brown color of the lower proportion of right hind leg and scapula region of the left front leg comes from Japanese Brown(parthenogenetic cell). However, this calf also died at day 2 after birth. Along with this parthology, the animla exhibited a blooding from heart and intestine, and umbilical vessels two times normal size. The number of placenta also small than that of normal(50 vs 60-80). We confirmed this calf is a chimera clone with parthenogenetic and fertilized cells by DNA analysis.

无琼脂包埋组的聚集率（53-70%）显著低于1.0%和1.2%琼脂包埋组（93-95%）（P<0.01）。然而，根据聚集胚胎的数量，各处理组的囊胚发育率没有差异。对通过将孤雌生殖胚胎注射到雄性受精胚胎中产生的聚集胚胎和通过整个孤雌生殖胚胎和受精胚胎聚集产生的聚集胚胎进行核型分析。聚集胚胎的核型分析结果显示，在同一样本中，53%的胚胎具有XX和XY染色体板，27%的胚胎具有XY染色体板，20%的胚胎具有XX染色体板。与未成熟胚相比，聚集胚的平均囊胚细胞数较低与全胚聚集组相比，差异极显著（P<0.01）（158*65 vs 202*32），11只受体中有6只在移植荷斯坦牛的聚集胚胎后被诊断为妊娠（用日本黑受精）和日本棕（孤雌生殖胚胎）：6头中有2头流产，获得7头小牛（3对双胞胎）。双胞胎雄性（死产）在妊娠第234天从受体分娩，其中在每头小牛中检测到XX和XY染色体板。在妊娠第261天分娩了检测到XX和XY染色体平板的单头雄性小牛。皮肤颜色也被用作嵌合体的证据。右后腿下部和左前腿肩胛骨区域的棕色来自日本棕色（孤雌生殖细胞）。然而，这头小牛也在出生后第2天死亡。沿着这一病理变化，阿尼玛表现出心脏和肠道出血，脐血管是正常血管的两倍。胎盘数量也较正常少（50个vs 60-80个）。通过DNA分析，我们证实这头小牛是一个具有孤雌生殖和受精细胞的嵌合体克隆。