拟南芥 AtTTG1基因的克隆及用于转化花生的 MAR 调控表达载体构建

摘要

Aflatoxin contamination has severely threatened the industry of peanut.We want to ap-ply gene engineering to change the structure of peanut's pod and testa to improve the ability of anti-A. flavus .According to the coding region cDNA of the Arabidopsis thaliana AtTTG 1 gene in the Gen-Bank, we designed the primers, and the AtTTG1 gene was cloned by reverse transcription PCR (RT-PCR).The result showed that the length of the cloned segment was 1026bp,it was same as the data of GenBank by compare.This segment coded 341 amino acids, the predicted protein was about 86.96 KDa, and its isoelectric point was about 4.85.The plant high efficient expression vector of pLMAR-S1 9-TTG1 was successfully constructed.It used pod and testa specific promoter S1 9 of peanut and the plant expression vector of pLMAR which was regulated by MAR sequence, and it was transformed in-to agrobacterium EHA105.This work aimed at laying a foundation for future transformation of pea-nut in order to obtaining transgenic peanut with high resistance to A.flavus .%黄曲霉毒素污染对花生产业危害巨大,通过基因工程改变花生果种皮结构以提高花生抗黄曲霉能力。根据 GenBank 中拟南芥 AtTTG1基因的 cDNA 编码区设计引物,通过 RT-PCR 克隆AtTTG1基因,结果显示,克隆获得的片段长1026bp,与基因库中数据比对相同,该片段编码341个氨基酸,预测其蛋白分子量为86.96KDa,等电点为4.85。结合实验室已获得的花生果种皮特异启动子 S19和 MAR 序列调控的植物表达载体 pLMAR,构建了植物高效表达载体 pLMAR-S19-TTG1,并将其导入根癌农杆菌 EHA105。为进一步对花生进行遗传转化,获得转基因高抗黄曲霉花生奠定基础。