先天性心脏病患者CHD7基因突变的研究

目的 筛查先天性心脏病(congenital heart fflsease,CHD)患者CHD7(chromodomsin helicase DNA-binding protein gene)基因的胚系突变,探讨其在先心病发生中的作用.方法 采集67例临床确诊的CHD患儿、100名正常对照的外周静脉血,提取白细胞基因组DNA,PCR扩增CHD7基因,变性高效液相色谱分析技术对PCR产物进行突变筛选,出现异常峰型的扩增片段进行DNA测序,明确突变位点和类型,并进一步结合病例对照和生物信息学分析探讨变异的功能意义.结果 67例先心病患者中共检出7种单碱基的替换,位于CHD7的不同内含子.其中,IVS11+127A＞G和WS12+21T＞G的等位基因频率均为0.0075,为罕见变异;而IVS2+34G＞A、IVS4+39C＞A、IVS12.5T＞C和IVS16+51C＞A等位基因频率为0.2635、0.2156、0.1505、0.3636,属于单核苷酸多态性;IVS12-5T＞C在CHD组的检出频率显著低于正常对照组(5.42%vs 9.57%,P＜0.05);而IVS14-35C＞G则仅见于CHD患儿.生物信息学分析显示,IVS12-5T＞C替换具有增强外显子剪切的效应.结论 CHD7基因单核苷酸多态性变异WS12-5T＞C对CHO的发生可能具有保护作用,而CHD7基因的突变并不是构成散发性先天性心脏病的主要原因.

Mutation analysis of the CHD7 gene in patients with congenital heart disease

Objective To investigate the germline mutations of the CHD7 gene and their roles in patients with congenital heart disease (CHD).Methods Genomic DNAs extracted from peripheral blood were subjected to screen mutations in CHD7 germ by denaturing high performance liquid chromatography (DHPLC) followed by DNA sequencing of aberrant peaks in 67 CHD patients and 100 healthy control.Case-control study and bioinformatie analysis were utilized to explore the potential functional roles of the variations detected.Resells Seven kinds of single nucleotide substitution were detected in the CHD patients in different introus of the CHD7 gene.Among them,IVS11+127A＞G and IVS12 +21T＞ G were rare variations and the allele frequencies of both were 0.0075;Mille IVS2 + 34G＞A,IVS4 + 39G＞A,IVS12-5T＞ C and IVS16 + 51C＞ A were the single nucleotide polymorphisms and the allele frequency was 0.2635,0.2156,0.1505 and 0.3636 respectively.The frequency of IVS12-5T＞ C in the CHD group was significantly lower than that in the control group (5.42% versus 9.57%,P ＜0.05).The variant of IVS14-35C＞G was only detected in patients with CHD.Bioinformatie analysis showed that IVS12-5T＞C might increase exon splicing ability comparing with the wild-type sequence.Conclusion The CHD7 germ mutation may not be the main reason for sporadic congenital heart disease,whereas the single nucleotide polymorphism of IVS12-5T＞C might play a protective role in the onset of this dis-ease.