肝豆状核变性基因8号内含子的研究

The study of sequence and structure of intron 8 in Wilson disease gene.

目的 分析Wilson病 (WD)基因 8号内含子的序列结构 ,探讨 8号内含子与WD发病的关系。方法 采用聚合酶链反应 (PCR)扩增 10个正常人、2 0例WD患者及其 12个亲属含WD基因 8号外显子 8号内含子 9号外显子 (8E 8I 9E)的DNA片段 ,进行直接测序。将测得的序列输入计算机进行分析。结果 ① 8号内含子的长度为70 3bp ,GC含量为 42 7%。②存在一个短串联重复序列和 7个正、反向重复序列。③在 132 137位核苷酸有一TATAbox ,其下游有一个编码 82个氨基酸的开放性阅读框架 (ORF)。④ 40 8、487位核苷酸存在G(A)多态。⑤ 5’端序列为GTAAC ,3’端序列为CCTAG ,剪接分枝点序列为TTTCGAT。结论 正常人、WD患者及其亲属的WD8号内含子序列和结构无差异 。

Objective To study the sequence and structure of intron 8 in WD gene in order to further understand the relationship between intron 8 and WD. Methods We utilized polymerase chain reaction (PCR) to the amplification of exon 8 intron 8 exon 9 which were then sequenced by a dideoxy chain termination methon in 10 normal controls and 32 members of 11 families(20 WD patients and 10 of their relative). The results were analyzed by the computer. Results The sequence of intron 8 was 703 bp with the G+C content of 42.7%. There were one short tandom repeats, 7 direct and inverted repeats in it. An open reading frame coded with 82aa was found at 323 base pairs of downstream of a TATAbox. There were two DNA polymorphisms at 408 and 487 nucleotides. The sequence analysis showed that the 5end has the sequence of 5 GTAAC, 3end has the sequence of CCTAG 3, and branchpoint of 5 TTTCGA 3.Conclusions The sequences and structures of intron 8 in WD familiess members are not different from normal controls. Our data suggest that the WD gene intron 8 might not play an important role in the pathogenesis of WD.