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目的运用微卫星(短串联重复序列,STR)结合的多重定量荧光PCR检测61例流产绒毛染色体数目异常,以评价它与传统经典的核型分析方法的关系。方法首先利用多重荧光定量PCR扩增48份无关个体DNA样本中的常见发生数目异常的染色体上19个STR位点和一个AMXY位点,扩增产物经毛细管电泳。计算出各位点的杂合度和正常杂合子峰面积比值范围。再对61例自然流产组织中这些位点进行扩增,判断染色体数目异常情况。染色体核型分析结果做为对照诊断标准。结果①核型分析成功率为65.6%;多重荧光定量PCR扩增成功率为98.3%。核型分析成功的40例样本,多重荧光定量PCR全部扩增成功,其中38例结果与核型分析结果相一致,以细胞遗传学作为诊断标准,诊断的符合率为95%。②运用核型分析方法,新鲜标本核型分析成功率为80.4%;不新鲜或者坏死标本的成功率为20%,P<0.001。运用STR-PCR方法,新鲜标本扩增成功率为100%;不新鲜或者坏死标本的成功率为93.3%,P=0.246。③AMXY和DXS8090、DXS8094联合诊断性别的成功率可达97.9%。结论结合STR的多重荧光定量PCR在诊断染色体数目异常中敏感性强,特异度高,可以做为细胞遗传学的补充。对于稽留流产和坏死变性的绒毛组织,STR-PCR也可以进行诊断。AMXY和DXS8090、DXS8094可用来进行性别诊断。
Objective To detect the abnormal chromosome number of 61 cases of abortion with multiplex quantitative fluorescent PCR combined with microsatellite (short tandem repeat, STR) to evaluate its relationship with traditional classical karyotyping methods. Methods A total of 19 STR loci and one AMXY locus on the chromosomes of 48 unrelated individual DNA samples were amplified by multiplex fluorescence quantitative PCR. The amplified products were analyzed by capillary electrophoresis. Calculate the heterozygosity of each locus and the range of the ratio of normal heterozygote peak area. Then 61 cases of spontaneous abortion tissue in these sites were amplified to determine the number of chromosome abnormalities. Chromosome karyotype analysis as a diagnostic criteria. Results ① The success rate of karyotype analysis was 65.6%; the success rate of multiplex PCR amplification was 98.3%. Forty cases of successful karyotype analysis were successfully amplified by multiplex PCR. Among them, 38 cases were consistent with the results of karyotype analysis. The coincidence rate of 95% was based on the diagnosis of cytogenetics. ② The success rate of karyotype analysis of fresh specimens was 80.4% by karyotyping method; the success rate of non-fresh specimens or necrotic specimens was 20%, P <0.001. The STR-PCR method showed that the success rate of fresh specimens was 100%, that of fresh specimens or necrotic specimens was 93.3%, P = 0.246. ③ AMXY and DXS8090, DXS8094 combined diagnosis of gender success rate up to 97.9%. Conclusion Multiplex quantitative PCR with STR is sensitive and specific in diagnosing chromosomal abnormalities and can be used as a supplement to cytogenetics. STR-PCR can also be used to diagnose chorionic vasculature with aborted abortion and necrosis. AMXY and DXS8090, DXS8094 can be used for gender diagnosis.