CAJ | 학술논문

为揭示PbCBL4基因在杜梨抗逆防御机制中的作用,采用PCR方法克隆了杜梨PbCBL4基因的cDNA和DNA序列,利用半定量RT-PCR和原核表达分析了该基因对非生物胁迫的转录响应。结果表明：PbCBL4基因的cDNA长度为639 bp,编码一个含212个氨基酸的蛋白；基因组DNA长度为1645 bp,包含8个外显子和7个内含子。 PbCBL4编码的蛋白具有植物类钙调磷酸酶B亚基蛋白结合Ca2+所必需的4个EF手型结构,1个典型的植物钙调磷酸酶A亚基结合位点。 PbCBL4与拟南芥AtCBL4亲缘关系最近,处于同一进化分支上。 PbCBL4基因在杜梨叶中为诱导型表达,对盐碱、干旱、渗透胁迫和ABA处理均存在转录响应,而在杜梨根中检测不到该基因的表达；将其转入大肠杆菌BL21(DE3)后,能够明显减轻NaCl、甘露醇和PEG6000对该菌株的生长抑制。综上所述, PbCBL4基因主要在叶片中参与了杜梨对非生物胁迫的防御机制。转入PbCBL4的重组大肠杆菌对盐胁迫和渗透胁迫的耐受能力均得到提高。
위게시PbCBL4기인재두리항역방어궤제중적작용,채용PCR방법극륭료두리PbCBL4기인적cDNA화DNA서렬,이용반정량RT-PCR화원핵표체분석료해기인대비생물협박적전록향응。결과표명：PbCBL4기인적cDNA장도위639 bp,편마일개함212개안기산적단백；기인조DNA장도위1645 bp,포함8개외현자화7개내함자。 PbCBL4편마적단백구유식물류개조린산매B아기단백결합Ca2+소필수적4개EF수형결구,1개전형적식물개조린산매A아기결합위점。 PbCBL4여의남개AtCBL4친연관계최근,처우동일진화분지상。 PbCBL4기인재두리협중위유도형표체,대염감、간한、삼투협박화ABA처리균존재전록향응,이재두리근중검측불도해기인적표체；장기전입대장간균BL21(DE3)후,능구명현감경NaCl、감로순화PEG6000대해균주적생장억제。종상소술, PbCBL4기인주요재협편중삼여료두리대비생물협박적방어궤제。전입PbCBL4적중조대장간균대염협박화삼투협박적내수능력균득도제고。
The objective of this study was to illuminate the role of PbCBL4 gene in the defense mechanism of birch-leaf pear ( Pyrus betulaefolia Bunge) . In this study, the cDNA and genomic DNA sequences of PbCBL4 gene from birch-leaf pear seedlings were isolated and cloned by EST database mining and PCR. PbCBL4 cDNA sequence is 639 bp in length encoding 212 amino acid residues. Its genomic DNA sequence is 1 645 bp in length consisting of 8 exons and 7 introns. PbCBL4 protein shares common structural features with CBLs from plants, which contain four EF-hand structure domains and one calcineurin A subunit binding sites. PbCBL4 and AtCBL4 from Arabidopsis belonged to the same branch in the CBL phylogenetic tree. Semi-quantitative RT-PCR analyses revealed that the mRNA abundance of PbCBL4 in birch-leaf pear leaves was responsive to salt, drought, osmotic stresses and ABA treatment. However, its expression was not detectable in roots of birch-leaf pear seedling with or without abiotic stress treatment. The inhibitory effects on Escheria coli strain BL21 ( DE3 ) growth caused by NaCl, mannitol or PEG6000 were significantly alleviated after PbCBL4 gene transformation. It is suggested that PbCBL4 gene might have played an role in defensive mechanism against abiotic stresses in the Pyrus betulaefolia leaves.