Basic research for the Wilson's disease gene therapy

以聚合酶链反应扩增的人威尔逊氏病cDNA片段为探针，发现LEC大鼠与对照组威尔森氏病基因存在限制性内切片段长度多态性。在LEC大鼠与WKAH大鼠的回交中发现该RFLP与肝功能障碍存在交叉连锁，提示LEC大鼠的肝功能障碍与人类威尔逊病的基因相同。提示LEC大鼠可作为肝豆状核变性基因治疗的有效动物模型。为了构建插入人Wilson病基因的逆转录病毒载体进行基因治疗，我们尝试从健康人肝脏cDNA文库中克隆该基因cDNA。然而，获得的cDNA克隆都没有编码整个编码区，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.