胰岛素样人参多肽基因的克隆

Chemical Synthesis and Cloning of Ginseng Peptide Gene

在对胰岛素样人参多肽进行了氨基酸顺序分析的基础上［１］，作者设计了该肽的基因并以化学法将基因分四段进行合成，合成好的片段经分离纯化后连接得小肽的完整基因．以ＰＵＣ９作为克隆载体，用ＢａｍＨＩ和ＳａｌＩ将其双酶切后与合成的小肽基因相连接得到重组质粒，作者以此重组质粒转化ＪＭ１０１菌株并以含氨苄青霉素和Ｘｇａｌ、ＩＰＴＧ的平板进行了筛选得到了白色的重组子菌落．此后作者通过原位杂交、Ｓｏｕｔｈｅｒｎ印迹杂交、双酶切电泳及顺序分析等重组子分析技术证明在得到的重组子中确有完全正确的小肽基因的克隆，这为进一步的表达打下了基础．

Upon the achievement of analyzing amino acid sequence of ginseng peptide, we synthesized the peptide gene chemically. Puc9 plasmid is taken as a cioing vector, after digestion with BamHI and SalI, it is ligated with the synthesized gene. The recombinant plasmid is used to tranformea JM101. The positive transformants of JM101were assayed by a few experiments: in situ hybridization, Southern blot, and physical map of the reoombinant plasmid. Finally, the sequencing result tells us the peptide gene is successfully cloned.