Use Of Protoplasts And Separate Cells In Plant Virus Research

Sander, Evamarie; Mertes, G.

doi:10.1016/s0065-3527(08)60410-8

Cited by 13 publications

(5 citation statements)

References 157 publications

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“…Their advantages include the stability of the cells, the ease with which the cultures can be handled, and the availability of the system without season-dependent physiological variations (110). However, the heterogeneous sizes of the plant cells, the asynchrony of cell growth, and the presence of a rigid cell wall often hampered efficient viral transfections.…”

Section: Protoplast Versus Other Systemsmentioning

confidence: 99%

Insights into the Single-Cell Reproduction Cycle of Members of the Family Bromoviridae : Lessons from the Use of Protoplast Systems

Sztuba-Solińska

Bujarski

2008

J Virol

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2The development of single-cell protoplast systems is certainly one of the milestones in the history of plant virology, allowing for the analysis of viral molecular processes at the cellular level. The use of plant cell-based systems in the study of the Bromoviridae family of multipartite single-stranded plant RNA viruses facilitated the discovery and dissection of viral processes engaged in the single-cell reproduction cycle: replication, transcription, protein synthesis, movement, virion assembly, and RNA recombination. This review summarizes the application of protoplast systems to the analysis of consecutive steps of the bromovirus life cycle, emphasizing their temporal and spatial patterns during virus multiplication.Our knowledge of viral infection at the individual cell level determines our understanding of the infection at the entire plant body level. The idea of applying single-cell systems in virology was introduced with the first attempts to transfect Escherichia coli cells with a T4 anti-E. coli bacteriophage (27). E. C. Cocking was the first (1960) to enzymatically isolate plant cells (19), which were then infected with Tobacco mosaic tobamovirus (TMV) (20). The ensuing pioneer work showed that the infection was synchronous and that the uptake of viral particles/RNAs by protoplasts was efficient enough to support virus replication (5, 118). The first viral infection of protoplasts involved the use of poly-L-ornithine (5), but the trials that followed exploited the fusogenic polymers (24), liposomes (28), or electroporation (78,129). By 1980, these procedures had been successfully employed to transfect protoplasts from more than six species of plants, including various members of the Bromoviridae family, e.g., Brome mosaic bromovirus (BMV; 80), Cucumber mosaic cucumovirus (CMV; 69), and Cowpea chlorotic mottle virus (CCMV; 130). The first attempts were based on the transfection of plant cells with whole-virus particles (104). Later on, viral RNA and its chemical modifications were used as the inoculum (62). Further advances in nucleic acid technology, especially the accessibility of infectious transcripts, have broadened the application of protoplast systems to the study of Bromoviridae.The Bromoviridae constitute one of the most important families of plant RNA viruses. They are distributed worldwide, they infect an extensive range of hosts, and some of them (e.g., CMV and Broad bean mottle bromovirus) are responsible for major crop epidemics (111, 63). The family consists of five genera named after their most representative members: Alfamovirus, Bromovirus, Cucumovirus, Ilarvirus, and Oleavirus. All these viruses possess tripartite, single-stranded, positive-sense RNA genomes (Fig. 1). RNA1 and RNA2 encode the RNAdependent RNA polymerase (RdRp) proteins 1a and 2a, respectively. The dicistronic RNA3 encodes the movement protein (MP) and coat protein (CP). The latter is translated from subgenomic RNA 4 (sgRNA4). Many of the family members, such as CMV, BMV, and Alfalfa mosaic alfamovirus (AMV), repres...

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Section: Protoplast Versus Other Systemsmentioning

confidence: 99%

Insights into the Single-Cell Reproduction Cycle of Members of the Family Bromoviridae : Lessons from the Use of Protoplast Systems

Sztuba-Solińska

Bujarski

2008

J Virol

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“…The first isolation of protoplasts was from tomato fruit, and the cells were inoculated with tobacco mosaic virus particles (Cocking, 1960;Cocking and Pojnar, 1969). Subsequent generalization of the technique for different starting materials (reviewed in Sander and Mertes, 1984), including cell suspension cultures, opened new avenues to study viruses at the cellular level. The use of this technique in viral research offers unique advantages.…”

Section: Commentary Background Informationmentioning

confidence: 99%

“…The use of cell suspension culture for protoplast isolation instead of plant tissues offers several advantages: (1) reproducibility of protoplast preparations with constant yield as the quality of cell suspension culture is less variable, (2) greater viability of the protoplasts, (3) reduced labor, and (4) more sterile conditions and thus less contamination by fungi or bacteria (reviewed in Sander and Mertes, 1984).…”

Section: Commentary Background Informationmentioning

confidence: 99%

Preparation and Electroporation of Oat Protoplasts from Cell Suspension Culture

et al. 2007

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Protoplasts provide a convenient system for introduction of nucleic acids into plant cells. Protoplasts allow rapid assays of gene expression and virus replication, with advantages for plant biology similar to those of cultured animal cells for the study of animal systems. Traditionally, preparation and handling of protoplasts has been as much art as science, requiring a special touch by the user. The purpose of this unit is to lay out in clear detail all the methods and nuances involved in protoplast preparation using a robust, reliable system that does not require skills beyond those expected of an unspecialized molecular biologist. Because dicots and monocots differ in many biological properties, and because different procedures may work better for different plants, separate units in this chapter are devoted to protoplast preparation from dicots (Arabidopsis, tobacco; refer to UNITS 16D.1 & 16D.4) and from a monocot (oat). This unit describes methods for preparation and transfection by electroporation of protoplasts derived from an oat suspension culture.

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“…Todas estas utilizações dos protoplastos mostram sua importância para a biologia em geral, mas na virologia, o uso de protoplastos tem um papel decisivo (COCKING, 1972;TAKEBE, 1975;SANDER & MERTES, 1984;WOOD, 1987 Nos últimos quinze anos uma grande quantidade de informações sobre vários aspectos da infecção de vírus de planta foi obtida através do uso de protoplastos (SHAW, 1985), possibilitando inclusive a co-infecção de protoplastos de um mesmo hospedeiro com diferentes vírus e viróides ou satélites para que se verifique as interações entre eles (MARCHOUX, 1988).…”

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Efeito da presença do vírus do “Vira-cabeça” do tomateiro e de ribavirina sobre a cinética de crescimento de protoplastos do mesófilo de fumo (>i<Nicotiana tabacum>/i< L.) em cultivo de suspensão e sobre isoformas de peroxidases, shiquimato desidrogenases e amilases

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Use Of Protoplasts And Separate Cells In Plant Virus Research

Cited by 13 publications

References 157 publications

Insights into the Single-Cell Reproduction Cycle of Members of the Family Bromoviridae : Lessons from the Use of Protoplast Systems

Insights into the Single-Cell Reproduction Cycle of Members of the Family Bromoviridae : Lessons from the Use of Protoplast Systems

Preparation and Electroporation of Oat Protoplasts from Cell Suspension Culture

Efeito da presença do vírus do “Vira-cabeça” do tomateiro e de ribavirina sobre a cinética de crescimento de protoplastos do mesófilo de fumo (>i<Nicotiana tabacum>/i< L.) em cultivo de suspensão e sobre isoformas de peroxidases, shiquimato desidrogenases e amilases

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