评价多重连接探针扩增法诊断染色体22q11.2微缺失结果研究

Objective To evaluate multiplex ligation-dependent probe amplification (MLPA) assay detection in analysis of chromosome 22q11.2 microdeletion. Methods Between March 2008 and September 2009, thirty-two patients including 10 males and 16 females aged between years (3.6±3.1) were selected and evaluated by history, physical examination and medical records. Of these patients, sixteen patients who were previous diagnostic as 22q11.2 microdeletion were in positive control group, the other 16 healthy children were in negative control group. All the patients were detected by MLPA and fluorescence in situ hybridization (FISH) for the presence of a 22q1 1.2 microdeletion after informed consent. Diagnostic efficacy was assessed by sensitivity, specificity and Kappa analysis. Results We have applied the two assays of detection of chromosome 22q11.2 microdeletion in 32 patients. Sixteen patients in positive control group were found to have a 22q11. 2 deletion and, with the deletion size of 3-Mb. However, as expected,chromosome 22q11.2 deletion was not found in negative control group. The MLPA results were in good agreement with that by FISH. Therefore, MLPA has high sensitivity and specificity. Conclusion MLPA is a rapid, reliable, high-throughput and relatively economical alternative to FISH technology for the diagnosis of 22q11.2 microdeletion. It can provide reliable and helpful information for clinical diagnosis of 22q11.2 microdeletion syndrome.     

评价多重连接探针扩增法诊断染色体22q11.2微缺失结果研究

Objective To evaluate multiplex ligation-dependent probe amplification (MLPA) assay detection in analysis of chromosome 22q11.2 microdeletion. Methods Between March 2008 and September 2009, thirty-two patients including 10 males and 16 females aged between years (3.6±3.1) were selected and evaluated by history, physical examination and medical records. Of these patients, sixteen patients who were previous diagnostic as 22q11.2 microdeletion were in positive control group, the other 16 healthy children were in negative control group. All the patients were detected by MLPA and fluorescence in situ hybridization (FISH) for the presence of a 22q1 1.2 microdeletion after informed consent. Diagnostic efficacy was assessed by sensitivity, specificity and Kappa analysis. Results We have applied the two assays of detection of chromosome 22q11.2 microdeletion in 32 patients. Sixteen patients in positive control group were found to have a 22q11. 2 deletion and, with the deletion size of 3-Mb. However, as expected,chromosome 22q11.2 deletion was not found in negative control group. The MLPA results were in good agreement with that by FISH. Therefore, MLPA has high sensitivity and specificity. Conclusion MLPA is a rapid, reliable, high-throughput and relatively economical alternative to FISH technology for the diagnosis of 22q11.2 microdeletion. It can provide reliable and helpful information for clinical diagnosis of 22q11.2 microdeletion syndrome.     

TBX1基因与染色体22q11.2微缺失综合征

染色体22q11·2微缺失综合征(22q11·2DS)是人类最常见的染色体微缺失综合征。TBX1基因作为一个T-BOX家族转录因子,其单拷贝缺失可能是22q11·2DS的主要原因之一,它对该综合征临床表征的出现可能有重要作用。TBX1在胚胎生长发育是胚胎咽部分节、咽弓和动脉弓形成、心脏流出道生长与排列及分隔等过程所必需的基因。TBX1受一系列调控基因调节,其本身也调控一系列基因。     

22q11.2微缺失综合征研究现况

22q11.2微缺失综合征是染色体22q11.21-q11.23缺失引起的遗传综合征,典型缺失区域内低重复序列(LCR22s)间的重组可能介导了缺失的发生。此综合征可表现为心脏、颅面、免疫等系统异常,已有研究对Tbx1、Ufd1L等相关基因进行探讨,现就其发病机制、临床特征、诊断与产前诊断作一综述。     

多重连接探针扩增检测Williams综合征微缺失

MLPA是一种简便快速检测缺失及重复突变的方法,该技术弥补了荧光原位杂交技术的不足,可用于实验室对Williams综合征的快速检测,具有一定临床诊断价值.     

22q11.2微缺失综合征产前临床表型的分析

目的探讨产前诊断的22q11.2微缺失综合征病例的不同临床表型,进一步提高对该疾病的认识。方法收集2016年1月至2018年12本院医学遗传中心因产前筛查高风险而就诊,应用染色体微阵列技术确诊的22q11.2微缺失综合征病例,分析其临床表现。结果14651例产前筛查高风险病例中22例(1.5‰)确诊为22q11.2微缺失综合征,其中心脏超声结构异常13例(59.09%),心脏结构异常合并腭裂1例(4.55%),足内翻4例(18.18%),肾脏超声异常3例(13.64%),胎儿发育迟缓并唐氏筛查高风险1例(4.55%);22例22q11.2微缺失综合征中经典型19例(66.36%),非经典型3例(13.64%)。结论本组22q11.2微缺失综合征病例以足内翻为突出临床表现,仅次于心脏结构异常,应予以重视。     

22q11.2微缺失综合征胎儿的产前诊断及家系分析

目的:分析22q11.2微缺失综合征胎儿的产前诊断、亲代验证及妊娠结局,为遗传咨询提供依据。方法:对低深度全基因组测序技术(copy number variation sequencing,CNV-Seq)或染色体微阵列分析(chromosomal microarray analysis,CMA)发现的6例22q11....     

人类染色体22q11.2微缺失综合征发病机制的研究进展

22q11缺失综合征（22qll deletion syndrome,22qllDS）是由染色22q11．21-q11．23缺失引起的遗传性综合征，其临床表现复杂，主要包括心脏、颅面、免疫等系统异常。22qll缺失产生的机制是缺失区域内低拷贝重复序列（10w-copy-repetitives，LCR22s）之间的不对称重组。本文对其临床表现、发病机制、候选基因克隆等方面的近年进展进行了综述。     

人类染色体22q11.2微缺失综合征发病机制的研究进展

22ql1缺失综合征(22q11 deletion syndrome,22q11DS)是由染色22ql1.21-ql1.23缺失引起的遗传性综合征，其临床表现复杂，主要包括心脏、颅面、免疫等系统异常。22q11缺失产生的机制是缺失区域内低拷贝重复序列(1ow-copy-repetitives，LCR22s)之间的不对称重组。本文对其临床表现、发病机制、候选基因克隆等方面的近年进展进行了综述。     

染色体微阵列产前诊断8q13.3微缺失一例

目的应用染色体微阵列分析(chromosome microarray analysis,CMA)技术对1例超声结构异常胎儿进行全基因组拷贝数变异(copy number variations,CNVs)检测,探讨CMA在超声结构异常胎儿产前诊断中的意义。方法应用常规G显带染色体核型分析胎儿及其父母的染色体核型,应用CMA技术分析胎儿及其父母的CNVs。结果G显带核型分析显示胎儿核型与母亲一致,为46,XN,t(8;11)(q21.2;q13)mat,父亲核型正常;父母CMA检测结果均未见异常;胎儿的检测结果为arr[GRCh37]8q13.3(71314082-73322915)×1,提示一条8号染色体的8q13.3区域发生2.00 Mb缺失。结论超声结构异常胎儿染色体核型分析检出的平衡易位,需借助CMA等技术进一步确定是否存在微缺失微重复。     

男性少精子患者染色体和Y染色体微缺失检查

目的探讨男性少精子患者与外周血染色体异常及Y染色体微缺失的关系和临床意义。方法对150例男性少精子门诊患者同时进行外周血染色体核型分析和Y染色体微缺失检测。对照组为40例已正常生育的男性。结果150例男性少精子患者中,染色体异常30例,异常率20%;Y染色体微缺失8例,缺失率5.3%。对照组40例正常男性均未发现染色体异常和Y染色体微缺失。结论染色体异常或Y染色体微缺失是引起男性少精子的重要原因之一,对临床上少精子患者有必要进行染色体核型分析和Y染色体微缺失检测。     

采用多重连接探针扩增技术检测智力低下儿童的基因缺陷

目的 探讨原因不明智力低下儿童的发病与染色体亚端粒基因重组间的关系.方法 采用多重连接探针扩增(multiplex ligation-dependent probe amplification,MLPA)技术检测30名原因不明的综合征性智力低下患儿的染色体亚端粒区域.结果 检测到5例患儿存在染色体亚端粒的基因缺失或重复突变,分别为4p缺失,21q重复,10p重复、4p缺失,15p重复,3p重复、9p缺失.结论 不明原因智力低下儿童的发病与染色体亚端粒基因重组密切相关.MLPA技术可以作为一种高效、特异的方法对智能障碍儿童进行基因缺陷筛查.     

染色体核型异常男性不育患者Y染色体微缺失分析

目的探讨染色体核型异常与Y染色体微缺失之间的关系。方法578例男性不育患者均来自2007年6月至2008年5月吉林省生殖医学研究所临床门诊。所有患者临床表现均为无精子症或严重少精子症。外周血淋巴细胞培养常规染色体标本制备,进行染色体核型分析。应用多重聚合酶链反应技术,采用无精子因子区9个序列标签位点对所有染色体异常的无精子症或严重少精子症患者进行Y染色体微缺失分析。结果578例遗传咨询患者中,检测出染色体核型异常患者62例,异常率为10.73%。其中包括无精子症或严重少精子症患者10例,占总样本1.73%。10例染色体核型异常患者检测出Y染色体微缺失2例,占20%。核型为46,XX/47,XX,＋del（Y）（q11）患者临床表现为睾丸小,无精症,Y染色体缺失位点为sY157、sY152、sY254、sY255;核型为45,X,-Y,-15,＋t（Y：15）（p？;q11）患者临床表现为特发性无精子症,缺失位点为sY143、sY254、sY255。结论涉及到Y染色体的染色体核型异常与AZF微缺失密切相关。     

应用多重连接依赖式探针扩增技术快速高通量诊断胎儿染色体非整倍体异常

Objective To assess the diagnostic value of multiplex ligation-dependent probe amplification (MLPA) for detection of common chromosome aneuploidy in amniotic fluid (AF) cells in order to obtain an accurate, rapid, cost-effective and high-throughput method in routine prenatal clinical practice.Methods The MLPA test was performed on 500 AF samples by using kit P095 and the results were obtained by using analysis software RH-MLPA-v511. The results were compared with that from fluorescence in situ hybridization (FISH) and traditional karyotyping (TK). The technical critical issues were analyzed in routine diagnostic application. Results The absolute specificity and sensitivity of the MLPA test to detect the aneuploidy were 100%. For the 500 AF samples, the success rate of the MLPA tests was 97%. Among them 92% were finished within three working days and 5% required more days for repeating. The test failure rate was 3%. The results confirmed that for the 38 detectable aneuploid samples,the probe reliability weighted mean ratio values were more than 4SD compared to normal diploids and the 2 suspected trisomy samples were more than 2SD. In this study, authors analyzed hybridization efficiencies of 8 probes for chromosome 21, and the presence of a trisomy was considered if at least 4 of the 8 probes gave probe ratio of ＞1.3. Conclusion The data suggested that MLPA is a rapid, simple and reliable method for large scale testing for aneuploidy of chromosomes 13, 18, 21, X, or Y in AF. The MLPA technology is complementary to AF culture and valuable for prenatal diagnosis.     

应用多重连接依赖式探针扩增技术快速高通量诊断胎儿染色体非整倍体异常

Objective To assess the diagnostic value of multiplex ligation-dependent probe amplification (MLPA) for detection of common chromosome aneuploidy in amniotic fluid (AF) cells in order to obtain an accurate, rapid, cost-effective and high-throughput method in routine prenatal clinical practice.Methods The MLPA test was performed on 500 AF samples by using kit P095 and the results were obtained by using analysis software RH-MLPA-v511. The results were compared with that from fluorescence in situ hybridization (FISH) and traditional karyotyping (TK). The technical critical issues were analyzed in routine diagnostic application. Results The absolute specificity and sensitivity of the MLPA test to detect the aneuploidy were 100%. For the 500 AF samples, the success rate of the MLPA tests was 97%. Among them 92% were finished within three working days and 5% required more days for repeating. The test failure rate was 3%. The results confirmed that for the 38 detectable aneuploid samples,the probe reliability weighted mean ratio values were more than 4SD compared to normal diploids and the 2 suspected trisomy samples were more than 2SD. In this study, authors analyzed hybridization efficiencies of 8 probes for chromosome 21, and the presence of a trisomy was considered if at least 4 of the 8 probes gave probe ratio of ＞1.3. Conclusion The data suggested that MLPA is a rapid, simple and reliable method for large scale testing for aneuploidy of chromosomes 13, 18, 21, X, or Y in AF. The MLPA technology is complementary to AF culture and valuable for prenatal diagnosis.     

多重链接探针扩增技术在染色体非整倍体畸变诊断中的应用

目的 探讨多重连接探针扩增(multiplexligation-dependent probe amplification,MLPA)技术在13、18、21、X、Y染色体非整倍体畸变诊断中的应用价值.方法 收集本院经染色体核型分析确诊包括有上述5种染色体数目异常及正常的样本共44份,其中外周血30份、胎儿脐带血10份、羊水4份,提取标本DNA,采用MLPA技术对样本染色体数目进行分析,并与染色体核型分析结果进行比对.结果 42例样本检测结果与染色体核型分析结果一致,1例染色体核型分析未能作出判断的标记染色体片段被识别为Y染色体片段,1例21-三体嵌合体未能做出明确判断,临床检出率97.7%(43/44).结论 MLPA技术通过单管反应同时检测40多个不同靶基因序列的拷贝数,具有高通量、特异、便捷及成本较低等特点,可应用于常见染色体非整倍体畸变的临床诊断和产前诊断.

Abstract：

Objective To investigate the application value of the multiplex ligation-dependent probe amplification (MLPA) technique in diagnosis and prenatal diagnosis of chromosomes 13, 18, 21, X and Yaneuploidy. Methods Forty-four cases including 30 peripheral blood samples, 10 fetal cord blood samples,and 4 amniotic fluid samples were collected in this study. DNA was isolated from the samples and detected by MLPA, followed by analyzing in ABI310 Genetic Analyzer. Analysis of copy number changes for chromosomes 13, 18, 21, X and Y was carried out with RH-MLPA-analysis software. The routine karyotype analyses were also done for all the samples. Results Of 44 samples, the results of 42 by MLPA method was consistent with that by chromosome karyotyping. Only one case with trisomy 21 chimerism was failed to reach conclusion. In addition, one case of mark chromosome segment was identified as Ychromosome segment by MLPA, while karyotyping failed to make judgment. The accurate rate of MLPA was 97. 7% (43/44). Conclusion The MLPA technique can simultaneously detect dozens of different target sequences and their copy number changes in a single reaction. It showed high specificity, good reproducibility, was fast and high-throughput. The MLPA technique can be applied to diagnosis and prenatal diagnosis of the common chromosomal aneuploidy.     

应用多重连接依赖式探针扩增技术快速高通量诊断胎儿染色体非整倍体异常

目的 评价多重连接依赖式探针扩增技术(multiplex ligation-dependent probe amplification,MLPA)在染色体非整倍体诊断中的应用价值,为我国羊水染色体诊断提供一种快速、特异、高通量的分子诊断手段.方法 应用MLPA技术检测了500份羊水标本,所有标本均进行荧光原位杂交(fluorescence in situ hybridization,FISH)技术检测和常规染色体核型分析,应用RH-MLPA-v511数据分析软件获得MLPA结果,比较MLPA技术与FISH和染色体核型分析结果的准确性,总结MLPA技术临床应用过程中的关键要点.结果 在500份羊水标本中,MLPA检测成功率97%.3个工作日完成结果的为92%,需重复检测的为5%,失败为3%.对染色体非整倍体异常检测敏感性和准确性100%.证实38例非整倍体病例探针信号比值＞正常二倍体4s,2例疑似三体结果＞2s.分析了21号染色体8条探针的杂交效率,21三体患者8条探针中平均4条探针比值＞1.3.结论 MLPA技术具有快速、特异、敏感、高通量、成本低等特点,可用于产前染色体非整倍体数目的快速检测,是传统染色体培养方法的补充,临床应用价值较高.

Abstract：

Objective To assess the diagnostic value of multiplex ligation-dependent probe amplification (MLPA) for detection of common chromosome aneuploidy in amniotic fluid (AF) cells in order to obtain an accurate, rapid, cost-effective and high-throughput method in routine prenatal clinical practice.Methods The MLPA test was performed on 500 AF samples by using kit P095 and the results were obtained by using analysis software RH-MLPA-v511. The results were compared with that from fluorescence in situ hybridization (FISH) and traditional karyotyping (TK). The technical critical issues were analyzed in routine diagnostic application. Results The absolute specificity and sensitivity of the MLPA test to detect the aneuploidy were 100%. For the 500 AF samples, the success rate of the MLPA tests was 97%. Among them 92% were finished within three working days and 5% required more days for repeating. The test failure rate was 3%. The results confirmed that for the 38 detectable aneuploid samples,the probe reliability weighted mean ratio values were more than 4SD compared to normal diploids and the 2 suspected trisomy samples were more than 2SD. In this study, authors analyzed hybridization efficiencies of 8 probes for chromosome 21, and the presence of a trisomy was considered if at least 4 of the 8 probes gave probe ratio of ＞1.3. Conclusion The data suggested that MLPA is a rapid, simple and reliable method for large scale testing for aneuploidy of chromosomes 13, 18, 21, X, or Y in AF. The MLPA technology is complementary to AF culture and valuable for prenatal diagnosis.