茶树细胞色素P450基因CYP710A1的克隆及差异表达特征分析

Cloning and Expression of CYP710A1 Gene of Cytochrome P450 in Camellia Sinensis

茶树细胞色素P450在茶树抵抗病虫害侵染过程中发挥重要作用。本研究利用NCBI中的茶树转录组数据库,获取了茶树细胞色素P450中的CYP710A1基因,通过RT-PCR及RACE技术克隆了该基因全长。通过分子生物学软件分析了该蛋白的氨基酸结构、蛋白质理化性质、蛋白功能域及系统进化树,并通过荧光定量技术(qRT-PCR)检测了该基因在不同叶片中的表达含量。结果表明,CYP710A1基因全长1911bp,包含1 506bp的编码序列(ORF),能够编码502个氨基酸,分子量大小为57.06kDa,理论等电点为7.64,包含一个P450蛋白结构域(Pfam domain)和一个跨膜结构域(transmembrane structure)。该基因与风铃椒C.baccatum亲缘关系较近,而与博落回M.cordata亲缘关系较远。同时,在单芽的表达量最低,在三叶中的表达量最高,三叶中的最高表达量是单芽的38.17倍。

Cytochrome P450 plays an important role in disease-resistance and insect infestation of tea plants.This study cloned the full cDNA sequence of cytochrome P450 CYP 710A1 gene of Camellia sinensis by RT-PCR and 5'/3'RACE based on the NCBI transcriptome database.The amino acid sequence,physicochemical properties,functional domain and phylogenetic tree of CYP 710A1 were determined using bioinformatics software,while the mRNA of the leaves at different positions on a plant compared applying qRT-PCR data.The results showed that the length of a complete cDNA of CYP 710A1 was 1,911 bp encoded 502 amino acids with a molecular weight of 57.06 kDa and the theoretical isoelectric point at 7.64.The gene consisted of a P450 Pfam domain with a transmembrane structure.A phylogenetic tree analysis showed that CYP 710A1 had a genetic relationship closest with Capsicum baccatum and farthest with Macleaya cordata.The qRT-PCR results indicated that the third leaves on a branch tip had the highest expression of CYP 710A1 gene,and the buds the lowest-with a 38.17-fold difference between them.