丝状蓝藻与大肠杆菌之间杂合质粒的构建与克隆

Construction and Clone of Hybrid-plasmid between Filamentous Cyanobacteria(Plectonema boryanum )and E.coli

丝状蓝藻ｐｌｅｃｔｏｎｅｍａｂｏｒｙａｎｕｍ的ｐＰｂ１经ＨｐａⅡ酶酶切与ＡｃｃⅠ酶酶切的ｐＵＣ１３构成杂合质粒，杂合质粒克隆到Ｅ．ｃｏｌｉＪＭ１０９、频率为３．４％．转化进Ｅ．ＣｏｌｉＪＭ１０９的杂合质粒，用ＥｃｏＲⅠ和ＨｐａⅠ酶切后得到证实．杂合质粒转化到新制备的Ｐ．ｂｏｒｙａｎｕｍ原生质球或丝状体后，再生的藻体表现出高于原来两倍以上的Ａｐ抗性．利用Ａｐ敏感的Ｅ．ｃｏｌｉ平板培养，证实了转化入藻体的质粒对Ａｐ具有高抗性．然而，获得的Ａｐ高抗株在后续培养中不能继代维持．

Hybrid-plasmid from Hpa Ⅱ-digested ppb_1and AccⅠ-cleaved pUC13 was cloned into E. coli JM109. The purified hybrid-plasmid was transformed backwardly into freshly prepared spheroplast or filament from P. boryanum. The transformed algal colonies displayed anti-ampicillin activity in a level up to two times higher concentration of Ap had been confirmed in plate culture with an AP sensitific E. coli strain. Nevertheless,the acquired Ap￣r can not withstand this level of AP on longer succesive culture.