两个先天性珊瑚状白内障家系CRYGD基因P23T突变的检测

背景先天性白内障的致病基因及临床表型具有明显的异质性，通过连锁分析进行基因定位，直接测序法筛选致病基因是目前常用的分析方法。目的对2个先天性珊瑚状白内障家系（CCl和CC2）进行致病基因研究。方法收集确诊为常染色体显性遗传的2个先天性珊瑚状白内障家系17名成员的外周静脉血各5ml，包括ll例患者、4名正常成员和2名配偶，提取基因组DNA。选取与已知常染色体显性遗传先天性白内障相关的位点进行基因组扫描，利用微卫星标记物进行PCR扩增并进行序列分析。采用两点法计算LOD值对致病基因进行连锁分析。采用直接测序法对候选致病基因以及3个单核苷酸多态性（sNP）位点（rs2305429，rs2305430，rs2242074）进行序列分析。利用SNP对2个家系的先证者进行单体型分析。结果CCl和CC2家系中的先证者裂隙灯下均可见双眼晶状体核中央混浊区呈珊瑚状，经两点法计算LOD值，家系CCl在微卫星位点D2S325获得的最大LOD值为3．28，而CC2家系在D2S325获得的最大LOD值为1．50，连锁分析结果支持2个家系均与位于2q的候选致病基因CRYGC和CRYGD连锁。基因序列分析发现2个珊瑚状白内障家系均携带CRYGD基因c．C70A．（P．P23T）突变体，而2个家系中的正常人及100名正常对照则无此基因突变。2个家系先证者携带不同类型的单体型结构。结论CRYGD基因C．C70A．（P．P23T）突变是导致2个不同祖先来源的先天性珊瑚状白内障家系CCl和CC2致病的丰要原因。

Recurrent P23T change in CRYGD gene in two Chinese families with congenital coralliform cataracts

Background Clinical and genetic heterogeneity of congenital cataract is well substantiated.Researchers often identify disease loci by linkage analysis and screen candidate gene by direct sequencing.Objective This study was to localize and identify the disease-causing genes for two Chinese families with congenital coralliform cataracts.Methods Two Chinese families(CC1 and CC2) with autosomal dominant inheritance congenital coralliform cataracts were ascertained and patients in the families underwent ophthalmological examination.Periphery blood samples were collected and DNA was extracted from 17 subjects including 11 cataract patients and 4 phenotype normal and 2 spouses.A linkage scan of genomic regions containing 25 known candidate genes was performed using 50 polymorphic microsatellite markers on genomic DNA from affected and unaffected family members and LOD scores were calculated.Candidate genes were sequenced and mutations were analyzed.Three single nucleotyde polymorphisms(SNP)(rs2305429,rs2305430,rs2242074) were sequenced and genotyped for the detect of the possibility of a common origin between CC1 and CC2.This study complied with the Declaration of Helsinki and was approved by Ethic Committee of Tianjin Eye Hospital.The informed consent was obtained from subjects and their guardian before the protocol.Results A significant LOD score of 3.28(θ=0) in family CC1 and a maximum LOD score of 1.50(θ=0) in family CC2 were both produced at the microsatellite marker D2S325 linked with CRYGD gene.Sequencing of CRYGD gene showed a heterozygous single base pair change c.70C＞A in exon2,predicting to result in a P23T amino acid change.The haplotypes of two probands in their respective families was quite distinct.Conclusion These results indicate that c.C70A(p.P23T) mutation in CRYGD gene is the underlyingmolecular pathogenesis of the two families with congenital coralliform cataracts,and this mutation occurs independently in these two families rather than descending from a common ancestor.