改良单细胞核固定和荧光原位杂交技术在卵裂期胚胎性别诊断中的应用

摘要

目的 建立一种快速便捷的单细胞核固定和荧光原位杂交(FISH)检测技术用于性别筛选,探究其可行性,同时分析植入前胚胎中非整倍体和嵌合体的发生情况.方法 解冻12位患者32枚胚胎,同时选择同期常规冻融胚胎移植周期23位患者51枚胚胎为囊胚培养对照组.激光打孔法活检卵裂期胚胎,每枚胚胎常规活检吸取2个卵裂球,活检后胚胎继续培养囊胚.改良吐温-20/HC1法游离和固定卵裂球细胞核,XY染色体计数探针和18号染色体计数探针同时孵育后荧光镜下观察结果.结果 32枚胚胎中,X、Y和18号染色体正常胚胎24枚,正常率75％;异常胚胎8枚,其中6例为嵌合体胚胎,嵌合体率18.8％.活检组与囊胚培养对照组的囊胚形成率和优质囊胚率比较,差异无统计学意义(P＞0.05).结论 本实验所建立的改良单细胞核固定和FISH技术能够快速有效地对卵裂期胚胎进行性别选择,对解决性染色体异常和性连锁性疾病性别诊断的生育障碍有一定价值.%Objective To optimize and evaluate the value of single blastomere nucleus fixation and fluorescence in situ hybridization (FISH) for gender screening of euploid and chimera.Methods The laser perforation method was used to extract two blastomeres for each embryo.Optimal Tween-20 / HCl method was performed to fix the nucleus.The X/Y chromosome probe and the 18-chromosome probe were also incubated simultaneously and the results were observed under the fluorescence microscope.Results Twenty-four embryos with X,Y and chromosome 18 normal embryos were found (normal rate 75％),and also 8 abnormal embryos,of which 6 were chimeric embryos with a chimerism rate of 18.8％.The blastocyst formation rate and high-quality blastocyst rate in the biopsy group were lower than those in the non-biopsy group (P ＞ 0.05,respectively).Conclusion Modified single blastomere nucleus fixation and FISH are useful for gender screening of cleavage stage embryos and valuable for solving the reproductive problem of sexual chromosomal abnormalities and sex-linked diseases.