ipt基因种子特异表达载体的构建及转基因大豆的获得

Construction of Seed-specific Expression Vector of ipt Gene and Regeneration of Transgenic Glycine max

细胞分裂素是重要的植物激素,它对植物的生长发育具有重要的调控作用。通过基因工程技术提高内源细胞分裂素在大豆种子中的含量,诱导营养物质向种子定向运输,可提高大豆的灌浆效率,为增加作物产量提供了新的研究方向。ipt基因编码的异戊烯转移酶是催化AMP转化为细胞分裂素的关键酶,将ipt基因与大豆种子发育中后期特异性表达的β-伴球蛋白α-亚基专一启动子(7ap)融合,构建了植物表达载体pCAMBIA33017αp-ipt-CaMV35 3'ULR,并通过农杆菌介导转化大豆。经草铵膦抗性筛选、PCR、GUS组织化学染色及Southern Blot检测,证实外源基因以单拷贝的形式整合进大豆基因组,共获得8株转基因大豆。

Cytokinin is an important phytohormone in the regulation of plant growth and development. Increas-ing the endogenous cytokinin content in soybean seed through genetic engineering can induce nutrients transport to seeds, improve the filling efficiency of soybean, and provide a new research direction for increasing crop yield. Isopentenyl-transferase encoded by ipt gene is the key enzyme in converting AMP to cytokinin. In order to study the effect of cytokinin in developing seed, we constructed a plant expression vector carrying cytokinin synthesis gene ipt coding sequence under the control of the seed-specific promoter（ 7α p） from Glycine max, and it was introduced into Glycine max mediated by Agrobacterium tumefaciens. It is confirmed that the exogenous genes were integrated into the soybean genome in the form of single copy by glufosinate ammonium resistance screening, PCR test, GUS assay and Southern blot analysis. At the last we obtained 8 lines transgenic soybean.