Construction of chimeric viruses based on pepper mild mottle virus using a modified Cre/loxP system |
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Authors: | YIN Yue-yan HUA Meng-ying ZHAO Kuang-jie WAN Qiong-lian BU Shan LU Yu-wen ZHENG Hong-ying RAO Shao-fei YAN Fei PENG Jie-jun CHEN Hai-ru CHEN Jian-ping |
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Affiliation: | 1. College of Plant Protection, Yunnan Agricultural University, Kunming 650201, P.R. China;2. Institute of Alpine Economic Plants, Yunnan Academy of Agricultural Sciences, Lijiang 674100, P.R. China;3. State Key Laboratory for Managing Biotic and Chemical Threats to the Quality and Safety of Agroproducts, Institute of Plant Virology, Ningbo University, Ningbo 315211, P.R. China;4. Longping Branch of Graduate College, Hunan University, Changsha 410125, P.R. China |
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Abstract: | Cre/loxP, a site-specific recombination system, has been widely used for various purposes, including chromosomal translocations, generation of marker-free transgenic plants, tissue-specific activation of a reporter gene and efficient heterologous gene expression in plants. However, stable or transient expression of Cre recombinase in plants can cause chlorosis or necrosis. Here, we describe a modified Cre/loxP recombination system using a DNA fragment flanked with loxP sites in the same orientation in which necrosis induced by Cre recombinase in Nicotiana benthamiana leaves was alleviated. The modified system was successfully used to create functional GFP-tagged pepper mild mottle virus (PMMoV) and a chimeric virus with coat protein (CP) substitution assembled from separate pro-vector modules. Our results provide a new strategy and flexible technique to construct chimeric virus and infectious clones for plant viruses with large genomes. |
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Keywords: | pepper mild mottle virus necrosis infectious cDNA clone chimeric virus |
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