蒺藜苜蓿二氢黄酮还原酶基因的克隆与原核表达

Cloning and prokaryotic expression of dihydroflavonol reductase gene from Medicargo Truncatula L.

以蒺藜苜蓿(Medicargo trunctula)幼果为试材,采用RT-PCR方法克隆到二氢黄酮还原酶(DFR)基因的cDNA片段.该片段全长1 018bp,与GenBank中已注册的DFR基因同源性为99.8%,编码334个氨基酸;构建DFR基因原核表达载体pETDFR,DFR在原核表达系统中得到了高效表达,获得预期大小(38.1ku)的酶蛋白分子.原核表达产物经SDS-PAGE分析表明,表达蛋白的分子质量与预期一致.

The full length DFR cDNA was amplified by reverse transcription-PCR（RT-PCR）from Medicargo truncatula L.fruitlets.The confirmed full length of this sequence was 1018 bp and shared a homology of 99.8% with registered Medicargo Truncatula L DFRgene in GenBank.The amplified gene encoded a protein of 334 amino acids.The prokaryotic expression vector was successfully constructed and named after pETDFR,the DFRgene was highly expressed in prokaryotic expression system.SDS-PAGE analysis showed the molecular mass of encoded protein was 38.1ku as expected.