椪柑柠檬苦素-UDP-葡萄糖基转移酶基因的克隆、分析及表达

目的克隆中药橘核主要来源品种椪柑Citrus reticulata的柠檬苦素-UDP-葡萄糖基转移酶(limonoid-UDP-glucosyl transferase gene,LGT)基因,并进行生物信息学及表达分析为橘核有效成分合成及调控机制研究提供基础。方法克隆获得LGT序列,通过生物信息学对该基因蛋白的特征进行分析,构建LGT与相关物种LGT的系统进化树,利用RT-PCR分析橘皮、橘肉以及橘核中LGT基因表达量,并进行分析和比较。结果测序所得椪柑LGT序列全长为1 530 bp,具有1 509 bp的完整开放阅读框,编码502个氨基酸。并对其蛋白二级、三级结构进行了分析和预测。基因表达分析结果发现椪柑LGT基因在橘核中表达量最低,其次为橘皮,表达量最高为橘肉。结论成功克隆、分析并表达了椪柑LGT基因,为橘核中柠檬苦素类物质合成及调控机制研究提供基础。

Cloning, analysis, and expression of limonoid UDP-glucosyl transferase gene in Citri Reticulatae Semen

Objective To cloning limonoid-UDP-glucosyl transferase (LGT) gene in Citrus reticulata, analyze the bioinformations of LGT, and compare the expression to provide the foundation for the composite and regulation mechanism of the active ingredient in Citri Reticulatae Semen. Methods LGT gene was cloned the protein characteristics were analyzed using bioinformatics and constructing phylogenetic tree. The expression of LGT in the different parts was analyzed using real time-PCR. Results A 1 530 bp LGT sequence in in C. reticulata, was obtained, which had a 1509 bp ORF and could encode 502 amino acids. And protein analysis and forecast of its secondary and tertiary structures were carried out. Gene expression analysis revealed the expression of citrus LGT was the lowest in semen, and was followed by skin, and the highest expression of fresh. Conclusion LGT gene of C. reticulata, and is successfully cloned, analyzed, and expressed a basis for the study of matter in the synthesis and regulation of limonoids is provided.