卵裂期优质胚胎的染色体嵌合体研究

Analysis of chromosomal mosaicism in good quality cleavage embryos

目的应用基于多次退火环状循环扩增(multiple annealing and looping-based amplification cycles,MALBAC)的单细胞测序技术分析卵裂期优质胚胎的嵌合体率及类型。方法收集接受卵胞浆内单精子显微注射治疗并生育正常后代的夫妇捐献的卵裂期冷冻优质胚胎,解冻,消化透明带,收集卵裂球,进行单细胞全基因组扩增及二代测序。结果共解冻胚胎23枚,收集184个卵裂球。175个(95.1%)卵裂球获得了测序结果,其中100个(57.1%)为非整倍体。在23枚胚胎中,3个(13.0%)为正常的二倍体,20个(87.0%)为嵌合体,其中包括5个(21.7%)非整倍体嵌合型、7个(30.4%)为二倍体-非整倍体嵌合型、5个(21.7%)为异常嵌合型、3个(13.0%)为无规律分离型。结论卵裂期优质胚胎中存在较高的染色体嵌合体率。复杂染色体异常或高比例异常细胞的嵌合体可能是影响胚胎发育潜能的重要因素。

Objective To apply single cell sequencing based on multiple annealing and looping amplification cycles(MALBAC)for the determination of the rate and type of mosaicisms of high-quality embryos at cleavage stage.Methods After thawing and removing of zona pellucida by enzymatic digestion,blastomeres were collected the high-quality embryos donated by couples whom had given birth to healthy offspring by intracytoplasmic sperm injection and embryo transfer.The whole genome of single cell was amplified and subjected to next generation sequencing.Results From a total of 23 embryos,184 blastomeres were collected.175(95.1%)of the blastomeres were successfully sequenced,of which 100(57.1%)were found to harbor chromosomal aneuploidies.Among the 23 embryos,3(13.0%)were diploid,20(87.0%)were mosaicisms,which included 5(21.7%)aneuploid mosaicisms,7(30.4%)diploid-aneuploid mosaicisms,5(21.7%)abnormal mosaicisms,and 3(13.0%)irregular segregations.Conclusion There is a high rate of chromosomal mosaicisms in high-quality cleavage embryos.Mosaicisms of complex chromosomal abnormality or with high proportion of abnormal cells may be an important factor affecting the potential of embryonic development.