Combined expression of chitinase and lipid transfer protein genes in transgenic carrot plants enhances resistance to foliar fungal pathogens
Combined expression of chitinase and lipid transfer protein genes in transgenic carrot plants enhances resistance to foliar fungal pathogens
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DOI：
10.1007/s00299-007-0368-x
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发表时间：
2007-09-01
影响因子：
6.2
通讯作者：
Punja, Z. K.
中科院分区：
文献类型：
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作者：
Jayaraj, J.;Punja, Z. K.
将由大麦几丁质酶 (chi-2) 和小麦脂质转移蛋白 (ltp) 组成的两个发病相关 (PR) 蛋白基因通过农杆菌介导的转化，使用草胺膦乙酰转移酶 (bar) 基因作为选择标记，单独或组合引入胡萝卜植物中。通过 PCR 和 RT-PCR 证实超过 75% 的再生植物对转基因呈阳性，并且对除草剂 Liberty 具有抗性（0.2%，v/v）。 Northern 分析和免疫印迹证实约 70% 的植物中存在转基因表达，但各个品系之间的表达水平存在差异。 Southern 分析揭示了每个转基因的一到三个拷贝。转基因植物接种了两种坏死营养型叶面真菌病原体（Alternaria radiicicola 和 Botrytis cinerea），当两种 PR 基因都表达时，与单基因转化体相比，转基因植物表现出显着更高的抗性。在表达这两种基因的植物中，葡萄孢属和链格孢感染的疾病减少水平分别为 95% 和 90%，而单基因转化体的疾病减少水平为 40-50%。 chi2和ltp基因可以组合部署在其他作物中，以显着增强对坏死性真菌病原体的抵抗力。
Two pathogenesis-related (PR) protein genes consisting of a barley chitinase (chi-2) and a wheat lipid-transfer-protein (ltp) were introduced singly and in combination into carrot plants via Agrobacterium-mediated transformation using the phosphinothricin acetyl transferase (bar) gene as a selectable marker. Over 75% of regenerated plants were confirmed to be positive for the transgenes by PCR and RT-PCR and were resistant to the herbicide Liberty (0.2%, v/v). Northern analysis and immunoblotting confirmed the expression of the transgenes in about 70% of the plants, with variable expression levels among individual lines. Southern analysis revealed from one to three copies of each transgene. Transgenic plants were inoculated with two necrotrophic foliar fungal pathogens, Alternaria radicicola and Botrytis cinerea, and showed significantly higher resistance when both PR genes were expressed compared to single-gene transformants. The level of disease reduction in plants expressing both genes was 95% for Botrytis and 90% for Alternaria infection compared to 40-50% for single-gene transformants. The chi2 and ltp genes could be deployed in combination in other crop plants to significantly enhance resistance to necrotrophic fungal pathogens.