CAJ | 학술논문

　　NADP依赖的苹果酸酶（NADP-ME）是C4光合途径关键酶。为了确定TaNADP-ME1基因的功能，利用重组技术将前期克隆到的TaNADP-ME1基因构建到原核表达载体pET32a，双酶切和PCR鉴定阳性克隆，CaCl2法转化大肠杆菌BL21（DE3）pLysS，IPTG诱导融合蛋白表达，Ni 2＋-NTA琼脂糖亲和层析柱纯化融合蛋白。成功获得了重组原核表达载体pETE1，TaNADP-ME1基因在BL21（DE3）pLysS中得到了融合表达，SDS-PAGE表明，融合蛋白分子量为80 kDa，并成功纯化到融合蛋白。
　　NADP의뢰적평과산매（NADP-ME）시C4광합도경관건매。위료학정TaNADP-ME1기인적공능，이용중조기술장전기극륭도적TaNADP-ME1기인구건도원핵표체재체pET32a，쌍매절화PCR감정양성극륭，CaCl2법전화대장간균BL21（DE3）pLysS，IPTG유도융합단백표체，Ni 2＋-NTA경지당친화층석주순화융합단백。성공획득료중조원핵표체재체pETE1，TaNADP-ME1기인재BL21（DE3）pLysS중득도료융합표체，SDS-PAGE표명，융합단백분자량위80 kDa，병성공순화도융합단백。
NADP-dependent malic enzyme(NADP-ME)is a key enzyme in C4 photosynthesis,the objective is to construct the TaNADP-ME1 gene into prokaryotic expression vector ,express fusion protein in E.coli and purify the fusion protein.The TaNADP-ME1 gene was constructed into expression vector pET 32a by recombination technolo-gy,recombination plasmid was identified by digestion with restriction enzymes and PCR amplification ,and trans-formed into BL21(DE3) pLysS by CaCl 2 method,fusion protein was induced by IPTG and purified by Ni agarose column.This research successfully acquired the recombination vector pETE 1,and TaNADP-ME1 gene was accurately expressed in BL21(DE3)pLysS,SDS-PAGE revealed that the molecular weight of purified fusion protein was about 80 kDa and successfully acquired the fusion protein .This study laid a good foundation for identification the gene function of TaNADP-ME1 gene.