黄曲霉漆酶基因 HIGS 载体的构建及对花生的转化

摘要

Aflatoxin contamination is a severe problem adversely affecting peanut food safety.Breeding a new peanut variety resistant to Aspergillus flavus is a lengthy process through conventional means,and the re-sults are far from satisfaction.Host -induced gene silencing (HIGS),as a novel technology for RNA interfer-ence,provides possibilities for creating disease -resistant crop plants.However,there is no reports on pea-nut.To breed new A.flavus -resistant peanut through genetic engineering,the HIGS vectors containing 2 lac-case genes,Lac1 and Lac2,were constructed and then were transformed into an A.flavus -susceptible peanut cultivar Yueyou 20.PCR -positive transgenic peanut seeds with Lac1 gene were obtained.According to the HIGS principle,if the transgenic peanut is infected by A.flavus,the laccase gene expression will be inhibited by dsRNA,thus the resistance of peanut to A.flavus will be induced.Based on this principle,whether the transgenic seeds had resistance to A.flavus infection would be identified next.The study layed foundations for exploring the relationship between laccase gene and A.flavus pathogenicity in peanut,and provided a new way for breeding peanut cultivars with A.flavus resistance.%黄曲霉毒素污染严重影响着花生食品安全。通过常规育种的方式培育抗黄曲霉花生新品种进展缓慢，效果也难如人意。HIGS （Host －Induced Gene Silencing，寄主诱导的转基因沉默）技术是一种新兴的RNA 干扰技术，为培育抗病植物提供了可能。但该技术在花生上的应用尚未见报道。为创造抗黄曲霉花生新品系，本研究利用该技术构建了两个黄曲霉漆酶基因（Lac1和 Lac2）的 HIGS 载体，并试图将其转化到易感黄曲霉的花生品种粤油20中，获得了转 Lac1基因的 PCR 阳性种子。根据 HIGS 的原理，黄曲霉侵染后，转基因花生中产生的 dsRNA 将抑制黄曲霉漆酶基因表达，从而使转基因花生对黄曲霉产生抗性。基于该原理，下一步将对转基因种子是否抗黄曲霉侵染进行鉴定。本研究为利用 HIGS 技术探索黄曲霉漆酶基因与黄曲霉致病性的关系奠定了基础，为培育抗黄曲霉花生品种提供了一条新思路。