RNA RECOMBINATION——INTRODUCING POLY(A) INTO Qβ PHAGE

Poly(A) molecule is inserted between two QβRNA molecules which have been partiallydegraded, one from the 3'-end and another from the 5'-end. E. coli HfrH is infected with thesemosaic RNA molecules and a few recombinated phages carrying poly(A) are obtained. Analysisof the RNA of 907 phage (one of these recombination phages) shows that the chain lengthof the poly(A) it contained is about 40.     

EXPRESSION OF SYNTHESIZED HIRUDIN GENE IN YEAST

Hirudin is a sort of polypeptides secreted from the salivary gland of medicinal leech. It is of potential importance in medicine. We designed and synthesized the hirudin gene based on the amino acid sequence of hirudin HV2, and expressed it using the yeast alpha factor expression system. The yeast strain stably carrying the hirudin expression plasmid was deduced by mutagenesis. After its cultivation in rich nutritious medium for 36—48 h, the hirudin expression product secreted into the culture fluid was 10—20 ATU/ml. The HPLCpure hirudin product could be obtained through a simpler purification procedure, its N-terminal amino acid sequence was identical with the natural product, and it showed potent anticoagulant and antithrombin activity. About 3000 ATU of pure hirudin witha specific activity of 6600 ATU/mg could be obtained from 500 ml of culture fluid.     

人工合成的水蛭素基因在酵母中的表达

本文按水蛭素HV2的氨基酸序列,设计并合成了水蛭素基因,并在酵母α因子表达系统中表达。通过诱变得到的稳定携带水蛭素表达质粒的酵母菌株,在丰富培养基中培养36—48 h后,可在培养液中测得10—20 ATU/ml的分泌的水蛭素表达产物。经过较简单的纯化步骤,得到HPLC纯的水蛭素,其N-端氨基酸序列与天然产物完全相同,并有强烈的抗凝血和抑制凝血酶的活力。从500 ml培养液中可得到约3000 ATU的纯水蛭素,其比活力为6600 ATU/mg。     

RNA重组实验——将poly(A)引入Qβ噬菌体

本文将poly(A)连接在两个部分降解的Qβ RNA分子中间,然后感染E.coli原生质体,得到了含有poly(A)的重组噬菌体,这些噬菌菌体可以感染E.coli Hfr H,并携带引入的poly(A)分子进行繁殖,对其中的一株——907噬菌体的RNA分析表明,它所携带的poly(A)链长约为40。