Generation of a Triple-Shuttling Vector and the Application in Plant Plus-Strand RNA Virus Infectious cDNA Clone Construction

Infectious cloning of plant viruses is a powerful tool for studying the reverse genetic manipulation of viral genes in virus–host plant interactions, contributing to a deeper understanding of the life history and pathogenesis of viruses. Yet, most of the infectious clones of RNA virus constructed in E. coli are unstable and toxic. Therefore, we modified the binary vector pCass4-Rz and constructed the ternary shuttle vector pCA4Y. The pCA4Y vector has a higher copy number in the E. coli than the conventional pCB301 vector, can obtain a high concentration of plasmid, and is economical and practical, so it is suitable for the construction of plant virus infectious clones in basic laboratories. The constructed vector can be directly extracted from yeast and transformed into Agrobacterium tumefaciens to avoid toxicity in E. coli. Taking advantage of the pCA4Y vector, we established a detailed large and multiple DNA HR-based cloning method in yeast using endogenous recombinase. We successfully constructed the Agrobacterium-based infectious cDNA clone of ReMV. This study provides a new choice for the construction of infectious viral clones.

[1]  Yuchen Zhou,et al.  Development of polyclonal antibodies-based serological methods for detection of the rehmannia mosaic virus in field plants , 2022, Frontiers in Sustainable Food Systems.

[2]  Sek-Man Wong,et al.  A plant virus hijacks phosphatidylinositol-3,5-bisphosphate to escape autophagic degradation in its insect vector , 2022, Autophagy.

[3]  Ting Yang,et al.  Construction of full-length cDNA infectious clones of Chilli veinal mottle virus. , 2022, Virus research.

[4]  Chuantao Lu,et al.  First Report of Tobacco Mild Green Mosaic Virus Infecting Rehmannia glutinosa in China. , 2022, Plant disease.

[5]  Xiaorong Tao,et al.  Rice stripe virus: Exploring Molecular Weapons in the Arsenal of a Negative-Sense RNA Virus. , 2021, Annual review of phytopathology.

[6]  Zhen He,et al.  Cucurbit chlorotic yellows virus infecting Rehmannia glutinosa was detected in China. , 2021, Plant disease.

[7]  Defu Wang,et al.  Complete Genome Sequencing and Infectious cDNA Clone Construction of Soybean Mosaic Virus Isolated from Shanxi , 2021, The plant pathology journal.

[8]  H. Wuriyanghan,et al.  Synthesis of Full-Length cDNA Infectious Clones of Soybean Mosaic Virus and Functional Identification of a Key Amino Acid in the Silencing Suppressor Hc-Pro , 2020, Viruses.

[9]  Bolan Yu,et al.  Novel genetically stable infectious clone for a Zika virus clinical isolate and identification of RNA elements essential for virus production. , 2018, Virus research.

[10]  Wei Zhang,et al.  Rapid Construction of Complex Plant RNA Virus Infectious cDNA Clones for Agroinfection Using a Yeast-E. coli-Agrobacterium Shuttle Vector , 2017, Viruses.

[11]  Y. Tong,et al.  Characterization of cis-Acting RNA Elements of Zika Virus by Using a Self-Splicing Ribozyme-Dependent Infectious Clone , 2017, Journal of Virology.

[12]  Xiaoying Li,et al.  Generation of stable infectious clones of plant viruses by using Rhizobium radiobacter for both cloning and inoculation , 2017, Virology.

[13]  Yongliang Zhang,et al.  The Barley stripe mosaic virus γb protein promotes chloroplast-targeted replication by enhancing unwinding of RNA duplexes , 2017, PLoS pathogens.

[14]  Xiaoying Li,et al.  Rapid Construction of Stable Infectious Full-Length cDNA Clone of Papaya Leaf Distortion Mosaic Virus Using In-Fusion Cloning , 2015, Viruses.

[15]  R. Blawid,et al.  Construction of an infectious clone of a plant RNA virus in a binary vector using one-step Gibson Assembly. , 2015, Journal of virological methods.

[16]  S. German-Retana,et al.  Gibson assembly: an easy way to clone potyviral full-length infectious cDNA clones expressing an ectopic VPg , 2015, Virology Journal.

[17]  J. Valkonen,et al.  Construction of an infectious cDNA clone and gene expression vector of Tobacco vein banding mosaic virus (genus Potyvirus). , 2012, Virus research.

[18]  H. Lecoq,et al.  A simple, rapid and efficient way to obtain infectious clones of potyviruses. , 2012, Journal of virological methods.

[19]  T. Candresse,et al.  Strategies to facilitate the development of uncloned or cloned infectious full-length viral cDNAs: Apple chlorotic leaf spot virus as a case study , 2011, Virology Journal.

[20]  M. Tepfer,et al.  A novel strategy for creating recombinant infectious RNA virus genomes. , 2004, Journal of virological methods.

[21]  A. Palloix,et al.  Mutations in potato virus Y genome-linked protein determine virulence toward recessive resistances in Capsicum annuum and Lycopersicon hirsutum. , 2004, Molecular plant-microbe interactions : MPMI.

[22]  J. García,et al.  Construction of a stable and highly infectious intron-containing cDNA clone of plum pox potyvirus and its use to infect plants by particle bombardment. , 2000, Virus research.

[23]  G. Stubbs,et al.  Tobacco Mosaic Virus: Pioneering Research for a Century , 1999, Plant Cell.

[24]  D. Baulcombe,et al.  Infectious in vivo and in vitro transcripts from a full-length cDNA clone of PVY-N605, a Swiss necrotic isolate of potato virus Y. , 1997, The Journal of general virology.

[25]  Robert H Levis,et al.  Infectious RNA transcripts from full-length dengue virus type 2 cDNA clones made in yeast , 1997, Journal of virology.

[26]  I. E. Johansen.  Intron insertion facilitates amplification of cloned virus cDNA in Escherichia coli while biological activity is reestablished after transcription in vivo. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[27]  J. Boyer,et al.  Infectious transcripts and cDNA clones of RNA viruses. , 1994, Virology.

[28]  M. Bray,et al.  Infectious RNA transcribed from stably cloned full-length cDNA of dengue type 4 virus. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[29]  S. MacFarlane,et al.  Construction and analysis of infectious transcripts synthesized from full-length cDNA clones of both genomic RNAs of pea early browning virus. , 1991, Virology.

[30]  I. Furusawa,et al.  Infectivity of plasmids containing brome mosaic virus cDNA linked to the cauliflower mosaic virus 35S RNA promoter. , 1991, The Journal of general virology.

[31]  K. Richards,et al.  In vitro synthesis of biologically active beet necrotic yellow vein virus RNA. , 1989, Virology.

[32]  Y. Shoyama,et al.  Identification of viruses infected in Rehmannia glutinosa Libosch. var purpurea Makino and effect of virus infection on root yield and iridoid glycoside contents , 1989, Plant Cell Reports.

[33]  L. Willmitzer,et al.  Storage of competent cells for Agrobacterium transformation. , 1988, Nucleic acids research.

[34]  P. Ahlquist,et al.  Multicomponent RNA plant virus infection derived from cloned viral cDNA. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[35]  Sanjeet Kumar,et al.  Virus diseases of peppers (Capsicum spp.) and their control. , 2014, Advances in virus research.

[36]  M. Jacquemond.  Cucumber mosaic virus. , 2012, Advances in virus research.

[37]  Tao Xiao-rong.  Construction of Agrobacterium-mediated Cucumber mosaic virus Infectious cDNA Clones and 2b Deletion Viral Vector , 2011 .

[38]  S. Chapman.  Construction of infectious clones for RNA viruses: TMV. , 2008, Methods in molecular biology.

[39]  A. Gibbs,et al.  Infectious eggplant mosaic tymovirus and ononis yellow mosaic tymovirus from cloned cDNA , 2005, Archives of Virology.

[40]  I. E. Johansen,et al.  Nucleotide sequence and infectious cDNA clone of the L1 isolate of Pea seed-borne mosaic potyvirus , 2001, Archives of Virology.

[41]  T. Candresse,et al.  Construction of full-length cDNA clones of lettuce mosaic virus (LMV) and the effects of intron-insertion on their viability in Escherichia coli and on their infectivity to plants , 1998, Archives of Virology.