肝素酶基因可变剪接体的克隆及鉴定

Cloning and Identification of Alternatively Spliced Variants of Human Heparanase Gene

目的:克隆肝素酶基因的可变剪接体并测序。方法:根据人肝素酶的cDNA序列设计引物,用RT-PCR方法从正常人外周血白细胞中扩增肝素酶基因的可变剪接体,构建至pGEM-T Easy载体中,转化大肠杆菌DH5α感受态细胞,筛选阳性克隆并进行序列测定。结果:获得了肝素酶基因的3种可变剪接体形式,即5号外显子缺失可变剪接体、6号外显子缺失可变剪接体、5和6号外显子缺失可变剪接体,其中后2种可变剪接体尚未报道。结论:克隆了肝素酶基因的3种可变剪接体,有助于研究各种肝素酶可变剪接体编码蛋白的结构和功能及其在肿瘤发生转移过程中的作用。

Objective： To clone and identify alternatively spliced variants of human heparanase gene.Methods： Alternatively spliced variants of human heparanase gene were amplified by RT-PCR,then ligated into pGEM-T Easy vector.After transformation,positive clones were identified by PCR and sequencing.Results： We obtained three alternatively spliced variants of human heparanase gene,lacked exon 5,exon 6,and exon 5 and 6 respectively.Conclusion： Heparanase gene have three alternatively spliced variants at least,thus investigation of structure and function of these variants is important for cancer occurrence and metastasis.