pSITE vectors for stable integration or transient expression of autofluorescent protein fusions in plants:: Probing Nicotiana benthamiana-virus interactions

被引:181
作者
Chakrabarty, Romit
Banerjee, Rituparna
Chung, Sang-Min
Farman, Mark
Citovsky, Vitaly
Hogenhout, Saskia A.
Tzfira, Tzvi
Goodin, Michael [1 ]
机构
[1] Univ Kentucky, Dept Plant Pathol, Lexington, KY 40546 USA
[2] Dongguk Univ, Dept Life Sci, Dongguk, South Korea
[3] SUNY Stony Brook, Dept Biochem & Cell Biol, Stony Brook, NY 11794 USA
[4] Ohio State Univ, Ohio Agr Res & Dev Ctr, Dept Entomol, Wooster, OH 44691 USA
[5] Univ Michigan, Dept Mol Cellular & Dev Biol, Ann Arbor, MI 48109 USA
关键词
agroinfiltration; confocal microscopy; nucleolus; SYNV; transformation;
D O I
10.1094/MPMI-20-7-0740
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Plant functional proteomics research is increasingly dependent upon vectors that facilitate high-throughput gene cloning and expression of fusions to autofluorescent proteins. Here, we describe the pSITE family of plasmids, a new set of Agrobacterium binary vectors, suitable for the stable integration or transient expression of various autofluorescent protein fusions in plant cells. The pSITE vectors permit single-step Gateway-mediated recombination cloning for construction of binary vectors that can be used directly in transient expression studies or for the selection of transgenic plants on media containing kanamycin. These vectors can be used to express native proteins or fusions to monmeric red fluorescent protein or the enhanced green fluorescent protein and its cyan and yellow-shifted spectral variants. We have validated the vectors for use in transient expression assays and for the generation of transgenic plants. Additionally, we have generated markers for fluorescent highlighting of actin filaments, chromatin, endoplasmic reticulum, and nucleoli. Finally, we show that pSITE vectors can be used for targeted gene expression in virus-infected cells, which should facilitate high-throughput characterization of protein dynamics in host-virus interactions.
引用
收藏
页码:740 / 750
页数:11
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