The Key Laboratory of Plant Resources Conservation and Germplasm Innovation in Mountainous Region (Ministry of Education), Institute of Agro-Bioengineering and College of Life Sciences, Guizhou University, Guiyang 550025, China.
The State Key Laboratory Breeding Base of Green Pesticide and Agricultural Bioengineering, Guizhou University, Guiyang 550025, China.
Int J Mol Sci. 2017 Nov 7;18(11):2361. doi: 10.3390/ijms18112361.
In this study, we cloned a new chitinase gene, , from Oliver () using rapid amplification of cDNA ends (RACE) technology and constructed an overexpression vector, pSH-35S-2, to introduce it into tobacco ( cv. Xanthi) Resistance to de Candolle (E.cichoracearum DC) and molecular mechanisms in the transgenic tobacco were determined by drop inoculation, spore counting, determination of physicochemical indicators, and analysis of gene expression. The chitinase activity and resistance to DC were significantly higher in the transgenic tobacco than in wild-type tobacco (p < 0.05). The activities of peroxidase (POD) and catalase (CAT), after inoculation with DC, were higher in the transgenic tobacco than in the wild-type. Conversely, the malondialdehyde (MDA) content was significantly lower in the transgenic tobacco than the wild-type before and after inoculation. In addition, our study also indicated that the resistance to DC might involve the salicylic acid (SA) and jasmonic acid (JA) pathways, because the expression levels of pathogenesis-related gene 1 () and () were significantly increased and decreased, respectively, after inoculation with DC. The present study supports the notion that and POD participate in resistance to DC in the transgenic tobacco plants.
在这项研究中，我们使用快速扩增 cDNA 末端（RACE）技术从 Oliver（）克隆了一个新的几丁质酶基因，并构建了一个过表达载体 pSH-35S-2，将其导入烟草（cv. Xanthi）。通过滴注接种、孢子计数、理化指标测定和基因表达分析，确定了转基因烟草对 E.cichoracearum DC 的抗性及其分子机制。与野生型烟草相比，转基因烟草的几丁质酶活性和对 DC 的抗性显著提高（p<0.05）。接种 DC 后，转基因烟草的过氧化物酶（POD）和过氧化氢酶（CAT）活性均高于野生型。相反，在接种前后，转基因烟草的丙二醛（MDA）含量均显著低于野生型。此外，我们的研究还表明，对 DC 的抗性可能涉及水杨酸（SA）和茉莉酸（JA）途径，因为接种 DC 后，病程相关基因 1（）和（）的表达水平分别显著增加和减少。本研究支持了几丁质酶和 POD 参与转基因烟草植株对 DC 抗性的观点。
