摘要
目的探讨高通量测序,即下一代测序(next-generation sequencing,NGS)技术在流产物遗传学分析和诊断中的应用价值。方法收集2017年1月至6月郑州大学第一附属医院的流产组织样本154例,分别采用NGS结合短串联重复序列(short tandem repeat,STR)和单核苷酸多态性芯片(single nucleotide polymorphism array,SNP-array)技术进行分析。比较2种平台的检测效果。采用χ^2检验或Fisher精确概率法对数据进行统计学分析。结果 (1)154例流产物样本中,检出染色体异常结果109例(70.7%)。109例染色体异常结果中,染色体数目异常52例(47.7%),染色体结构异常49例(45.0%),嵌合体6例(5.5%),单亲二倍体(uniparental disomy,UPD)2例(1.8%)。52例染色体数目异常样本共包括45例染色体非整倍体和7例染色体多倍体。染色体异常比例前3位依次为45,X(27.0%,14/52),22-三体(9.6%,5/52)和16-三体(7.7%,4/52)。49例染色体结构异常样本共携带67个拷贝数变异(copy number variation,CNV),其中致病性CNV13个(19.4%),临床意义不明的CNV24个(35.8%),良性CNV30个(44.8%)。在检出的致病性CNV中,2个包含微缺失/微重复综合征。(2)SNP-array技术对152例样本检测成功,2例(1.3%)因基因组DNA量<200 ng而检测失败。NGS技术对154例样本全部检测成功,且在SNP-array检测失败的2例中检出染色体异常。SNP-array和NGS技术检出的染色体异常比例差异无统计学意义[70.4%(107/152)与67.5%(104/154),χ^2=0.293,P=0.588]。(3)NGS和SNP-array技术各检出6例(均为3.9%)相同嵌合体和45例(分别为29.2%和29.6%)相同染色体非整倍体,差异均无统计学意义。SNP-array技术检出3例染色体多倍体(均为69,XXX)和UPD 2例,而NGS均未检出。但结合STR技术,NGS也成功检出了这3例染色体多倍体。(4)SNP-array检出的47例结构异常样本共携带CNV53个,NGS技术检出的49例结构异常样本共携带CNV67个。(5)当基因组长度分别为100~<500、500~<1 000和≥1 000 kb时,与SNP-array技术相比,NGS技术多检出的CNV分别为10、3和1个。结论NGS技术可以检出SNP-array技术检出的染色体非整倍体和嵌合体,对基因组DNA含量要求更低,还能够发现SNP-array技术不能检出的CNV。结合STR技术,NGS技术也可以有效检出染色体多倍体。NGS技术可以作为流产物遗传学诊断中的有效检测方法。
Objective To investigate the value of next-generation sequencing (NGS) technique for genetic analysis of spontaneous abortion.Methods From January to June 2017, 154 patients who visited the First Affiliated Hospital of Zhengzhou University for spontaneous abortion were enrolled. All abortion tissue samples were analyzed by both NGS combined with short tandem repeat (STR) and single nucleotide polymorphism array (SNP-array). Results of the two methods were compared by Chi-square or Fisher's exact test.Results (1) Chromosomal abnormalities were detected in 109 of the 154 cases (70.7%), including 52 (47.7%) of numerical chromosomal abnormalities, 49 (45.0%) of structural chromosomal abnormalities, six (5.5%) of mosaicism, and two (1.8%) of uniparental disomy (UPD). In those 52 cases of numerical chromosome abnormalities, there were 45 of chromosome aneuploidy and seven of polyploidy. The top three numerical chromosomal abnormalities were 45,X (27.0%, 14/52), trisomy 22 (9.6%, 5/52) and trisomy 16 (7.7%, 4/52). Forty-nine structural abnormality cases carried 67 copy number variations (CNV), including 13 pathogenic CNV (pCNV, 19.4%), 24 variants of unknown clinical significance (35.8%) and 30 benign CNV (44.8%). In those 13 pCNVs, two were responsible for microdeletion and microduplication syndromes. (2) SNP-array was successful in 152 cases, but failed in two (1.3%) due to genomic DNA <200 ng. However, NGS technology was successful in all 154 cases and identified chromosomal abnormalities in the two cases that SNP-array had failed. No statistically significant difference was shown in the detection rate of chromosomal abnormalities between SNP-array and NGS technology [70.4% (107/152) vs 67.5% (104/154), χ^2=0.293, P=0.588]. (3) No significant difference in the detection of chromosome aneuploidy (six cases in each group, 3.9% vs 3.9%) and mosaicism (45 cases in each group, 29.2% vs 29.6%) was found between NGS technology and SNP-array. Three cases of polyploidy (69, XXX) and two of UPD were identified by SNP-array, but not by NGS. When combined with STR, NGS was able to detect all three cases of polyploidy (69, XXX). (4) Forty-seven structural abnormality cases detected by SNP-array carried 53 CNVs, and 49 detected by NGS carried 67 CNVs. (5) NGS detected ten, three and one more CNVs than SNP-array did when the genome lengths were 100-<500, 500-<1 000 and ≥1 000 kb, respectively.Conclusions NGS can be used to detect chromosomal aneuploidy and mosaicism that can be identified by SNP-array with fewer limitations on total amount of genome. Moreover, CNVs that fail to be identified by SNP-array can also be detected by NGS. When combined with STR, NGS can effectively detect chromosomal polyploidy. Therefore, NGS could be a potential genetic analysis method for spontaneous abortion and of importance for genetic counseling.
作者
郭依琳
顾茂胜
王莉
屈素真
薛淑文
王海舰
玄兆伶
孔祥东
Guo Yilin;Gu Maosheng;Wang Li;Qu Suzhen;Xue Shuwen;Wang Haitian;Xuan Zhaoling;Kong Xiangdong(Center of Prenatal Diagnosis,First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China;Center of Medical Genetics,Xuzhou Maternal and Child Health Hospital,Xuzhou 211009,China;Annoroad Gene Technology Co.Ltd,Beijing 100176,China)
出处
《中华围产医学杂志》
CAS
CSCD
北大核心
2018年第12期808-816,共9页
Chinese Journal of Perinatal Medicine
关键词
流产
自然
高通量核苷酸序列分析
染色体畸变
多态性
单核苷酸
Abortion,spontaneous
High-throughput nucleotide sequencing
Chromosome aberrations
Polymorphism,single nucleotide
