摘要
利用聚合酶链式反应（ＰＣＲ）技术，从真养产碱杆菌Ａｌｃａｌｉｇｅｎｅｓ ｅｕｔｒｏｐｈｕｓ Ｈ１６ 染色体ＤＮＡ 中扩增并克隆了调控聚－β－羟基丁酸（ｐｏｌｙ－β－ｈｙｄｒｏｘｙｂｕｔｙｒａｔｅ， ＰＨＢ）生物合成的两个关键酶基因：依赖ＮＡＤＰＨ 的乙酰乙酰ＣｏＡ 还原酶基因（ｐｈｂＢ）和ＰＨＢ合成酶基因（ｐｈｂＣ）。限制性内切酶图谱和核苷酸序列分析证实了克隆结果，并表明所克隆的基因与国外报道的有很高的同源性。经过基因拼接，构建了块茎特异性表达的高等植物表达载体ｐＰＳＡＧＢ（嵌合ｐｈｂＢ）、ｐＢＩＢＧＣ（嵌合ｐｈｂＣ）和ｐＰＳＡＧＣＢ（嵌合ｐｈｂＢ和ｐｈｂＣ双基因）
NADPH dependent acetoacetyl CoA reductase gene (phbB) and poly β hydroxybutyrate (PHB) synthase gene (phbC) for biosynthesis of PHB were amplified and cloned from chromosomal DNA of Alcaligenes eutrophus H16 using PCR. The restriction maps and sequencing results show that both phbB and phbC have been cloned. It was found that the two genes cloned were highly homologous in DNA sequences to those being reported abroad. By DNA processing, the authors have constructed three tuber specific plant expression vectors: pPSAGB (containing phbB), pBIBGC (containing phbC) and pPSAGCB (containing both phbB and phbC).
