【摘要】 用固相亚磷酰胺法(solid-phase phosphoramidite method)合成了人α-心钠素(简称α-hANP)的全基因。合成的α-hANP基因全长为96bp,包括编码α-hANP的结构基因、基因5＇端的谷氨酸(作为表达产物用内肽酶Glu-C专一裂解的位点)密码子GAA、基因3＇端的终止信号TGA及基因两端的接头部分。基因分7个寡聚核苷酸片段在DNA合成仪上合成,然后一次性酶促连接成为完整的α—hANP双链基因。化学合成的基因克隆到M13载体上,经分子杂交鉴定筛选出的α-hANP基因克隆株,用双脱氧链终止法进行序列分析,证明核苷酸顺序正确。更多还原

【Abstract】 A total gene for a-human atrial natriuretic peptide(a-hANP) has been chemically synthesized by solid-phase phosphoramidite method. The synthetic gene consisted of 96 bp contains the structural gene encoding a-hANP, a glutamic acid codon GAA at 5’end of the gene for enzymatic cleavage of expressed product by endoproteinase Glu-C, a stop signal TGA at 3’ end of the structural gene and two restriction site at both ends of the gene. The synthesis of the gene involved enzymatic joining oligonucleotides to form DNA duplexes. The synthetic gene was cloned into vector M13.The phage progenies bearing a-hANP gene were identified by dot blotting and the gene sequence were analysed by dideoxynucleotide chain termination method.更多还原