一个血小板无力症家系的基因诊断及产前诊断

Mutation screening and prenatal diagnosis of a pedigree with Glanzmann＇s thrombasthenia

目的对一个血小板无力症家系进行基因诊断及产前诊断。方法应用PCR、逆转录-PCR、测序、限制性酶切分析及AT克隆分析等技术，从表达水平及基因组水平对该家系先证者进行仃GA2B与JTGB3基因诊断；进一步取母亲4个半月的胎儿羊水进行ITGA2B的产前诊断。结果先证者ITGA2B基因编码区第477～506的99个碱基发生缺失，导致密码子160～192缺失。进一步检测基因组DNA，确定为第4外显子的c．374C〉G点突变和第4内含子的＋5位G〉C点突变（IVY4＋5G〉C）复合突变体。其母亲为c．374C〉G点突变的携带者，其父亲为IVS-4＋5G〉C点突变的携带者，其中后者为剪切结构位点的点突变，而前者为菲剪切结构位点突变，但二者导致了相同的RNA剪切效果。产前诊断结果显示胎儿未遗传其父母的突变，为两个位点均正常的健康个体。先证者及其父母ITGB3基因未见异常。结论C．374C〉G和IVS-4＋5G〉C两个突变均为人类突变数据库和血小板无力症数据库未记载的新突变，二者导致相同的mRNA剪切异常，为该家系的疾病的致病原因。

Objective Mutation screening was performed in a pedigree of Glanzmann＇ thrombasthenia （GT） and prenatal diagnosis was performed. Methods In this study, reverse transcription- PCR-sequencing and PCR-sequencing, as well as restriction fragment length polymorphism（RFLP） and A/ T-cloned sequencing, were used to screen the ITGA2B and ITGB3 mutation in a pedigree with Glanzmann＇s thrombasthenia in the RNA and DNA level. Prenatal diagnosis was performed for this pedigree. Results Deletion of 99 bps was found in the eDNA of the patient in the pedigree, leading to deletion of 33 codons （from codon 160 to 192）. After genomic analysis, the patient was found to be a compound heterozygote of c. 374C〉G mutation and intron 4（IVS-4） -I-5 G〉C mutation. The two mutations were inherited from the parents. IVS-4 ＋5 G〉C mutation was a point mutation in the splice site, while c. 374C〉G mutation was out of the splice site. But both of them resulted in the same splice pattern in RNA. The two mutations were novel mutations which have not been reported in Human Gene Mutation Database（HGMD） and the mutation data base of Glanzmann＇s thrombasthenia. The results of ITGB3 gene screening is normal in the proband and his parents. Conclusion Two novel mutation, c. 374C〉G and IVS-4 ＋5G〉C were found in this study, which might be the cause of GT in the pedigree.