应用变性高效液相色谱技术检测三个遗传性多发性外生骨疣家系的EXT1与EXT2基因突变

目的建立一种应用变性高效液相色谱(denaturing high performance liquid chromatograph,DHPLC)技术检测遗传性多发性外生骨疣(hereditary multiple exostosis,EXT)基因突变的新方法,并研究3个EXT家系的基因突变情况。方法扩增EXT致病基因的所有外显子区及部分内含子与外显子交界区,联合连锁分析、DHPLC筛查及测序的方法进行分析。结果3个EXT家系中,共检测到两处已知EXT2基因的剪接位点突变IVS2 1G>A、IVS7 1G>T,一处28C>A变异和一处EXT1基因的IVS4 66G>A变异。结论EXT2基因的2个剪接位点突变分别是引起相应EXT家系的致病原因,应用DHPLC技术检测遗传性疾病基因变突是一种自动化、高通量、灵敏、快速且简便经济的好方法。

EXT1 and EXT2 mutation identified by denaturing high performance liquid chromatograph in three families with hereditary multiple exostoses

Objective To develop a new denaturing high performance liquid chromatograph(DHPLC)-based method to screen patients with EXT gene mutation and to study the gene mutation in three families with multiple exostoses.Methods All the exons of EXT gene,including the intro-exon boundaries,were amplified by PCR.Linkage analysis and DHPLC screening were carried out to identify the mutations.DNA sequencing was used to confirm the mutations.Results Two known splice site mutations,IVS2 1 G>A and IVS7 1 G>T,and two SNPs have been detected in EXT2 or EXT1 gene.Conclusion The transversions of IVS2 1 G>A and IVS7 1 G>T in EXT2 gene are suggested to be the disease-causing mutations and the DHPLC is a high throughout,sensitive,simple,quick,economical method to screen gene mutation in hereditary multiple exostosis.