【摘要】 为研究小麦与条锈菌互作过程中β-1,3-葡聚糖酶基因的功能,根据小麦β-1,3-葡聚糖酶cDNA全长序列设计合成引物,通过RT-PCR方法获得小麦β-1,3-葡聚糖酶基因编码区,将其克隆至pGEM T-easy Vector,经HindⅢ和BamHⅠ双酶切回收目的片断,并将其定向克隆到pET-32a(+)Vector中构建表达载体GLU-pET-32a(+),转化E.coliBL21(DE3),用IPTG进行诱导表达。结果表明,小麦β-1,3-葡聚糖酶基因在大肠杆菌中得到了高效表达,获得了分子量约为49 ku的融合蛋白,表达量约占菌体总蛋白的19%。最佳诱导条件为:0.3 mmol/LIPTG,37℃诱导4 h。更多还原

【Abstract】 To determine the function of β-1,3-glucanase in wheat to stripe rust resistance,the encoding region of a wheat β-1,3glucanase gene induced by Puccinia striiformis Westend f.sp tritici Eriks was engineered via reverse transcription polymerase chain reaction(RT-PCR).The amplified fragments were first cloned into pGEM T-easy Vector,after restriction analysis the gene was transferred to an E.coli strain BL 21(DE3) expression behind T7 promotor-pET-32a(+) system.The recombinant gave rise to a 49 ku fusion protein in response to the IPTG induction;under the best inducement condition(37 ℃,0.3 mmol/L IPTG,4 h) its content was about 19% of the total cell protein by Gene Genius Bio Imaging System.更多还原