串联的人胸腺素α1基因在番茄中的高效表达

 【目的】提高免疫活性多肽人胸腺素α1（Tα1）在番茄中的表达量。【方法】根据植物偏爱密码子对Tα1基因进行优化，并将其顺式串联成四联体，同时在单体基因间插入肠激酶剪切位点基因序列，以便表达的融合蛋白可以被肠激酶切割成单体。利用农杆菌介导法将单体Tα1基因和四联体Tα1基因分别转化进入番茄。【结果】两个基因转化番茄后分别得到卡那霉素抗性再生植株19株和10株，PCR和Southern杂交检测证明，部分番茄基因组中整合了外源基因。经过Western和ELISA分析，串联的Tα1基因已经在番茄果实中表达，表达量最高相当于20.2 μg Tα1?g-1果实鲜重，高于单体基因的表达量。【结论】基因串联的方式提高了Tα1基因在番茄中的表达量。

High Level Expression of Human Thymosin αl Concatemer in Transgenic Tomato Plants

【Objective】 To improve the expression of immunoreactive thymosin alpha 1(Tα1) protein in tomato plants.【Method】 The Tα1 gene was designed and synthesized according to the plant codon usage bias, then created a novel Tα1 concatemer that four copies of Tα1 gene were arranged end-to-end in tandem. Recognition site sequence of enterokinase was inserted between single Tα1 gene, so that fusion protein could be cut by enterokinase. Subsequently, Tα1 gene and Tα1 concatemer were respectively introduced into tomato via Agrobacterium tumefaciens mediated transformation. 【Result】 After the two genes transformation, 19 and 10 regenerated tomato plants resistant to kanamycin were obtained. Some of transgenic tomato plants were further confirmed by polymerase chain reaction (PCR) and Southern blot. Western blot analysis showed the expression of Tα1 concatemer protein in transgenic tomato fruits. The enzyme-linked immunosorbent assay (ELISA) analysis showed that the content of the Tα1 concatemer protein was equal to 20.2 μg Tα1?g-1 fresh weight in mature tomato fruit, which was higher than the expression of Tα1 gene. 【Conclusion】 The expression of Tα1 protein had increased in tomato by gene repeats.