肿瘤血管抑制肽alphastatin基因克隆及序列分析

目的克隆获得编码具有生物学活性的肿瘤血管抑制肽alphastatin基因。方法采用非对称引物／模板法，依据基因库提供的alphastatin基因序列，设计合成alphastatin基因的引物／模板链，利用聚合酶链反应（PCR）法，从DNA中扩增出人肿瘤血管抑制肽alphastatin基因片段；将获得的基因片段插入质粒载体pGEM．TEasy中，转化到大肠杆菌DH5ct后挑选阳性克隆，利用限制性内切酶酶切鉴定重组质粒。结果经质粒DNA酶切分析及序列测定，获得了人肿瘤血管抑制肽alphastatin基因片段序列。结论首次体外克隆获得了人肿瘤血管抑制肽alphastatin基因，为肿瘤血管抑制肽alphastatin的基因治疗奠定了基础。

Cloning and sequencing of antianiogenic peptide alphastatin gene

Objective Molecular cloning and sequencing of the antianiogenic peptide alphastatin gene. netlmds According to sequence of mature alphastatin supplied by GenBank, a double strand cDNA of mature alphastatin was chemically synthesized by using asymmetrical primer/template method which included a restrict enzyme side on the two extremmation and cloned into the pGEM-T Easy. pGEM-T/A1 was confirmed by endonuclease analysis and sequencing. Results Alphastatin gene was successfully synthesized chemically in vitro and cloned into plasmid pGEM-T Easy. The sequencing result of positive clone was identical to what sequenced and supplied by GenBank. Conclusion By cloning the alphastatin gene, the sequencing result of positive clone is identical to what sequenced and supplied by GenBank. It has paved the way for further study on gene therapy of tumor.