Basic research for the Wilson's disease gene therapy

用聚合酶链反应（PCR）扩增的Wilson病cDNA片段作探针，发现LEC和对照WKAH大鼠Wilson病基因的限制性片段长度多态性。发现该RFLP与LEC和WKAH大鼠之间的回交中的肝脏疾病显示交叉连锁，表明LEC大鼠中的肝脏疾病是由于与人类Wilson病的情况相同的基因。为了构建人Wilson病基因的逆转录病毒载体，我们从健康人肝cDNA文库中克隆了人Wilson病基因cDNA。然而，获得的cDNA克隆均不编码整个编码区，可能是由于该基因的编码区相对较长。我们正在重新筛选cDNA文库，以获得编码整个编码区的cDNA。

Restriction fragment length polymoyphism on the Wilson's disease gene betweem LEC and control WKAH rats was found by using a human Wilson's disease cDNA fragment amplified with polymerase chain reaction as the probe. This RFLP was found to show a cross-linkage with the hepatic disorder in back-crosses between LEC and WKAH rats, indicating that the hepatic disorder in LEC rats is due to the same gene as the case of human Wilson's disease. This result suggests that LEC rats are usuful animal model for the Wilson's disease gene therapy.In order to construct a retro virus vector inserted the human Wilson's disease gene for the gene therapy, we tried cloning of this gene cDNA from the cDNA library made from the healthy human liver. However, cDNA clones obtained all did not encode the whole coding region, possibly due to the relatively long coding region of this gene. We are under re-screening the cDNA library to obtain cDNA encoding the whole coding region.