CAJ | 학술논문

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 ITGA2B与 ITGB3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(IVS-4+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,而前者为非剪切结构位点突变,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.
목적 대일개혈소판무력증가계진행기인진단급산전진단.방법 응용PCR、역전록-PCR、측서、한제성매절분석급AT극륭분석등기술,종표체수평급기인조수평대해가계선증자진행 ITGA2B여 ITGB3기인진단;진일보취모친4개반월적태인양수진행 ITGA2B적산전진단.결과 선증자 ITGA2B기인편마구제477～506적99개감기발생결실,도치밀마자160～192결실.진일보검측기인조DNA,학정위제4외현자적c.374C＞G점돌변화제4내함자적+5위G＞C점돌변(IVS-4+5G>C)복합돌변체.기모친위c.374C>G점돌변적휴대자,기부친위IVS-4+5G>C점돌변적휴대자,기중후자위전절결구위점적점돌변,이전자위비전절결구위점돌변,단이자도치료상동적RNA전절효과.산전진단결과현시태인미유전기부모적돌변,위량개위점균정상적건강개체.선증자급기부모 ITGB3기인미견이상.결론 c.374C>G화IVS-4 +5G>C량개돌변균위인류돌변수거고화혈소판무력증수거고미기재적신돌변,이자도치상동적mRNA전절이상,위해가계적질병적치병원인.
Objective Mutation screening was performed in a pedigree of Glanzmann's 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 cDNA 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) +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.