原发性开角型青光眼患者TIGR基因突变筛选、序列分析

Screening and sequence analysis of TIGR gene mutation in patients with primary open angle glaucoma

目的:观察原发性开角型青光眼(primaryopenangleglaucoma,POAG)患者小梁网糖皮质激素诱导反应蛋白(trabecularmeshworkinducedglucocorticoidresponseprotein,TIGR)基因的突变情况。方法:实验于2001-01/2005-12在哈尔滨医科大学和暨南大学完成。应用聚合酶链式反应方法扩增90例原发性开角型青光眼患者和30例正常对照组的小梁网糖皮质激素诱导反应蛋白基因3个外显子的片段,应用单链构像多态筛选可能突变的聚合酶链反应产物。将聚合酶链反应产物,以EcorⅠ酶切鉴定重组质粒,然后测序。结果:①用单链构像多态方法筛选的5例原发性开角型青光眼患者在小梁网糖皮质激素诱导反应蛋白基因第3外显子中部(第6对引物)片段出现单链带型异常,余均未见异常;正常对照组未见异常。基因的3个外显子片段,包含各外显子的7对引物。②克隆测序的3例样品,在388密码子位置出现由GAT突变为AAT,氨基酸由天冬氨酸替换为天冬酰胺,即Asp388Asn。结论:原发性开角型青光眼患者的发病可能与小梁网糖皮质激素诱导反应蛋白基因突变有关。

AIM： To study the gene mutation of trabecular meshwork induced glucocorticoid response protein （TIGR） in patients with primary open angle glaucoma （POAG）. METHODS： The experiment was conducted in Harbin Medical University and Jinan University from January 2001 to December 2005. The polymerase chain reaction （PCR） was used to amplify the fragments of three exons in TIGR of 90 patients with POAG and 30 normal controls. The PCR amplification products of mutation was screened by single stranded conformation polymorphism （SSCP）. The PCR amplification products were identified of the construction plasmids by Ecorlendonuclease and the sequence was detected. RESULTS： （1）Single strand band abnormality was found in the middle fragments of the third exon （the sixth pair of primer） of TIGR gene in 5 patients with POAG with SSCP screening, while no abnormality was detected in other parts, Patients in the normal control group were normal. Three exons fragments of gene included 7 pairs of primer of all exons,（2） A ‘GAT＇ to ‘AAT＇ transition was found in the position of 388 codon in three samples that cloned and sequenced, and the amino acids was taken place by asparagine from aspartate, i.e. Asp 338 Asn. CONCLUSION： The pathogenesis of patients with POAG may be related with TIGR gene mutation.