论文部分内容阅读
以红皮梨‘奥冠’为试材,采用RT-PCR结合RACE技术获得1个MYB基因,命名为PyMYBa。该基因开放读码框共714 bp,编码237个氨基酸。PyMYBa分子量27.4 kD,等电点8.78。氨基酸序列分析显示,在其N端具有保守的R2R3-MYB结构域,R3-MYB结构域含有bHLH结合基序。进化树分析表明,PyMYBa与花青苷调控MYB转录因子的同源性很高。基因表达结果表明,PyMYBa在梨叶、花、果皮中表达量明显高于果肉,果皮中的表达量与花青苷调控基因PyMYB10相比,无显著差异。遮光及MeJA处理后果皮中PyMYBa、PyMYB10的表达量变化与花青苷合成量变化趋势相似,推测PyMYBa为梨花青苷合成中重要的正向调控因子。通过原核表达试验获得了该基因的重组蛋白,SDS-PAGE电泳检测结果与预期蛋白分子量一致。
Taking the red pear “Aoluan” as test material, one MYB gene was obtained by RT-PCR combined with RACE technology and named as PyMYBa. The gene was 714 bp in open reading frame and encoded 237 amino acids. PyMYBa has a molecular weight of 27.4 kD and an isoelectric point of 8.78. Amino acid sequence analysis revealed a conserved R2R3-MYB domain at its N-terminus and a R3-MYB domain with a bHLH binding motif. Phylogenetic tree analysis showed that PyMYBa and anthocyanin have high homology to MYB transcription factors. The results of gene expression showed that the expression level of PyMYBa in pears leaves, flowers and peels was significantly higher than that in pulp, and the expression level in peels was not significantly different from that in PyMYB10. The changes of the expression of PyMYBa and PyMYB10 in shading and MeJA-treated peel were similar to those of anthocyanin synthesis, suggesting that PyMYBa was an important positive regulator in the synthesis of pear anthocyanin. The recombinant protein of the gene was obtained by prokaryotic expression assay. The result of SDS-PAGE electrophoresis was consistent with the expected protein molecular weight.