聚球藻7942高效泌氨突变种的获得及其泌氨、谷氨酰胺合成酶活性、光合和生长

Establishment of a Highly Efficient Ammonia Secreting Mutant of Synechococcus sp. PCC 7942 and the Glutamine Synthetase Activity, Photosynthesis and Growth in Its Immobilized Cells

将含有Ａｎａｂａｅｎａｓｐ．ＰＣＣ７１２０反义ｇｌｎＡ基因片段的穿梭表达质粒ｐＤＣ－ＡＴＧＳ转化单细胞蓝藻聚球藻Ｓｙｎｅ－ｃｈｏｃｏｃｃｕｓ ｓｐ．ＰＣＣ７９４２，通过同源重组，外源ＤＮＡ定位整合到染色体上。经过抗菌素筛选，获得一种高效泌氨的Ｓｙｎｅｃhｏｃｏｃｃｕｓ ｓｐ．７９４２突变种。将此突变种固定化在聚氨脂泡沫中后，定量测定其谷氨酰胺合成酶（ＧＳ）活性。结果表明，固定化后的突变藻培养９ｄ后泌氨活性比自由生活的野生藻高１５６倍，ＧＳ活性降低７３．６％；其生长速度与同条件下野生藻相近，７７Ｋ荧光光谱表明突变种固定化后光系统Ⅱ活性提高４４％。

A recombinate plasmid pDC-ATGS was constructed, which contained the antisense fragment ofglaA gene from Anabaena sp. PCC 7120 and transformed the unicellular cyanobacterium Synechococcus sp.PCC 7942. The foreign DNA was inserted into the site of glnaA locus of the chromosome through the homologous recombination. By using neomyisin, a highly efficient ammonia secretion mutant was selected. After immobilized, the cells of the mutant within polyurethane (PU) foams, glutamine synthesize (GS) and NH4+ se- cretory activity of GS, and its growth and photosynthesis were measured. It was shown that NH4+ secretion ofthe immobilized mutant was enhanced 156 folds which was much higher than that of free-living cells of the wildtype. The activity of GS was decreased by 73. 6%. Growth of the mutant was the same as that of the wildtype. The activity of photosystem II in the immobilized mutant cells increased by 44% with 77 K fluorescencespectroum measurement.