慢生大豆根瘤菌(Bradyrhizobium japonicum)聚羟丁酸合成酶基因(phbC)的克隆和序列测定

CLONING AND SEQUENSING OF POLY-3-HYDROXYBUTYRATE SYNTHASE GENE (phbC) IN BRADYRHIZOBIUM JAPONICUM USDA110 STRAIN

以苜蓿根瘤菌Rm10 2 1的 phaC基因突变体菌株Rm1114 4 (phaC ::Tn5 - 2 33)为受体菌 ,通过功能互补 ,成功地从构建的Bradyrhizobium japonicum USDA110基因文库中 ,筛查到能与Rm1114 4互补 ,使之恢复在以乙酰乙酸为唯一碳源的M9培养基 (M9-AA)平板上 5d形成明显可见菌落 ,以及在MOPS平板上形成粘液型菌落的表型的重组粘粒 pDC2 ;经证实 ,该粘粒带有 phbC基因 .完成了该基因的全序列测定并已在GenBank登记 ,登记号为AY0 775 80 .B .japonicumphbC基因由 180 3碱基对组成 ,GC含量 6 1.8% ,AT含量 38.2 % ;编码 6 0 0个氨基酸 ,Mr=6 6 .95× 10 3 .图 3表 3参

By using the phaC gene mutant strain Rm11144 (phaC::Tn5-233) of Rhizobium meliloti Rm1021 as recipient and through functional complementation test, a recombinant cosmid pDC2 was screened from Bradyrhizobium japonicum USDA110 genomic library. The plasmid pDC2 could complement with Rm11144 strain, made it to show obvious colonies within 5 days on M9 medium with acetoacetate as sole carbon source and gave mucoid colonies on MOPS solid medium. The entire sequence of phbC gene was sequenced and deposited in GenBank at the accession number of AT077580. The phbC gene consistes of 1803 base pairs with GC 61.8%, AT 38.2% and encodes a 600 amino- acid protein with the M r of 66.95 ×10 3. Fig 3, Tab 3, Ref 13