Geminivirus vectors for high-level expression of foreign proteins in plant cells

Tsafrir Leket-Mor, Yong Sun Moon, Kenneth E. Palmer, Hugh Mason

Research output: Contribution to journalArticle

62 Citations (Scopus)

Abstract

Bean yellow dwarf virus (BeYDV) is a monopartite geminivirus that can infect dicotyledonous plants. We have developed a high-level expression system that utilizes elements of the replication machinery of this single-stranded DNA virus. The replication initiator protein (Rep) mediates release and replication of a replicon from a DNA construct ("LSL vector") that contains an expression cassette for a gene of interest flanked by cis-acting elements of the virus. We used tobacco NT1 cells and biolistic delivery of plasmid DNA for evaluation of replication and expression of reporter genes contained within an LSL vector. By codelivery of a GUS reporter-LSL vector and a Rep-supplying vector, we obtained up to 40-fold increase in expression levels compared to delivery of the reporter-LSL vectors alone. High-copy replication of the LSL vector was correlated with enhanced expression of GUS. Rep expression using a whole BeYDV clone, a cauliflower mosaic virus 35S promoter driving either genomic rep or an intron-deleted rep gene, or 35S-rep contained in the LSL vector all achieved efficient replication and enhancement of GUS expression. We anticipate that this system can be adapted for use in transgenic plants or plant cell cultures with appropriately regulated expression of Rep, with the potential to greatly increase yield of recombinant proteins.

Original languageEnglish (US)
Pages (from-to)430-437
Number of pages8
JournalBiotechnology and Bioengineering
Volume81
Issue number4
DOIs
StatePublished - Feb 20 2003

Fingerprint

Geminiviridae
Plant Cells
Viruses
Proteins
Biolistics
Caulimovirus
DNA
Genes
Replicon
DNA Viruses
Single-Stranded DNA
Genetically Modified Plants
DNA Replication
Reporter Genes
Recombinant Proteins
Introns
Plant cell culture
Tobacco
Plasmids
Clone Cells

Keywords

  • Bean yellow dwarf virus
  • Geminivirus
  • Plant gene expression
  • Recombinant protein
  • Transgenic plant

ASJC Scopus subject areas

  • Biotechnology
  • Microbiology

Cite this

Geminivirus vectors for high-level expression of foreign proteins in plant cells. / Leket-Mor, Tsafrir; Moon, Yong Sun; Palmer, Kenneth E.; Mason, Hugh.

In: Biotechnology and Bioengineering, Vol. 81, No. 4, 20.02.2003, p. 430-437.

Research output: Contribution to journalArticle

@article{4e65524d41d144e4bce1097d86bd9aca,
title = "Geminivirus vectors for high-level expression of foreign proteins in plant cells",
abstract = "Bean yellow dwarf virus (BeYDV) is a monopartite geminivirus that can infect dicotyledonous plants. We have developed a high-level expression system that utilizes elements of the replication machinery of this single-stranded DNA virus. The replication initiator protein (Rep) mediates release and replication of a replicon from a DNA construct ({"}LSL vector{"}) that contains an expression cassette for a gene of interest flanked by cis-acting elements of the virus. We used tobacco NT1 cells and biolistic delivery of plasmid DNA for evaluation of replication and expression of reporter genes contained within an LSL vector. By codelivery of a GUS reporter-LSL vector and a Rep-supplying vector, we obtained up to 40-fold increase in expression levels compared to delivery of the reporter-LSL vectors alone. High-copy replication of the LSL vector was correlated with enhanced expression of GUS. Rep expression using a whole BeYDV clone, a cauliflower mosaic virus 35S promoter driving either genomic rep or an intron-deleted rep gene, or 35S-rep contained in the LSL vector all achieved efficient replication and enhancement of GUS expression. We anticipate that this system can be adapted for use in transgenic plants or plant cell cultures with appropriately regulated expression of Rep, with the potential to greatly increase yield of recombinant proteins.",
keywords = "Bean yellow dwarf virus, Geminivirus, Plant gene expression, Recombinant protein, Transgenic plant",
author = "Tsafrir Leket-Mor and Moon, {Yong Sun} and Palmer, {Kenneth E.} and Hugh Mason",
year = "2003",
month = "2",
day = "20",
doi = "10.1002/bit.10483",
language = "English (US)",
volume = "81",
pages = "430--437",
journal = "Biotechnology and Bioengineering",
issn = "0006-3592",
publisher = "Wiley-VCH Verlag",
number = "4",

}

TY - JOUR

T1 - Geminivirus vectors for high-level expression of foreign proteins in plant cells

AU - Leket-Mor, Tsafrir

AU - Moon, Yong Sun

AU - Palmer, Kenneth E.

AU - Mason, Hugh

PY - 2003/2/20

Y1 - 2003/2/20

N2 - Bean yellow dwarf virus (BeYDV) is a monopartite geminivirus that can infect dicotyledonous plants. We have developed a high-level expression system that utilizes elements of the replication machinery of this single-stranded DNA virus. The replication initiator protein (Rep) mediates release and replication of a replicon from a DNA construct ("LSL vector") that contains an expression cassette for a gene of interest flanked by cis-acting elements of the virus. We used tobacco NT1 cells and biolistic delivery of plasmid DNA for evaluation of replication and expression of reporter genes contained within an LSL vector. By codelivery of a GUS reporter-LSL vector and a Rep-supplying vector, we obtained up to 40-fold increase in expression levels compared to delivery of the reporter-LSL vectors alone. High-copy replication of the LSL vector was correlated with enhanced expression of GUS. Rep expression using a whole BeYDV clone, a cauliflower mosaic virus 35S promoter driving either genomic rep or an intron-deleted rep gene, or 35S-rep contained in the LSL vector all achieved efficient replication and enhancement of GUS expression. We anticipate that this system can be adapted for use in transgenic plants or plant cell cultures with appropriately regulated expression of Rep, with the potential to greatly increase yield of recombinant proteins.

AB - Bean yellow dwarf virus (BeYDV) is a monopartite geminivirus that can infect dicotyledonous plants. We have developed a high-level expression system that utilizes elements of the replication machinery of this single-stranded DNA virus. The replication initiator protein (Rep) mediates release and replication of a replicon from a DNA construct ("LSL vector") that contains an expression cassette for a gene of interest flanked by cis-acting elements of the virus. We used tobacco NT1 cells and biolistic delivery of plasmid DNA for evaluation of replication and expression of reporter genes contained within an LSL vector. By codelivery of a GUS reporter-LSL vector and a Rep-supplying vector, we obtained up to 40-fold increase in expression levels compared to delivery of the reporter-LSL vectors alone. High-copy replication of the LSL vector was correlated with enhanced expression of GUS. Rep expression using a whole BeYDV clone, a cauliflower mosaic virus 35S promoter driving either genomic rep or an intron-deleted rep gene, or 35S-rep contained in the LSL vector all achieved efficient replication and enhancement of GUS expression. We anticipate that this system can be adapted for use in transgenic plants or plant cell cultures with appropriately regulated expression of Rep, with the potential to greatly increase yield of recombinant proteins.

KW - Bean yellow dwarf virus

KW - Geminivirus

KW - Plant gene expression

KW - Recombinant protein

KW - Transgenic plant

UR - http://www.scopus.com/inward/record.url?scp=0037456181&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0037456181&partnerID=8YFLogxK

U2 - 10.1002/bit.10483

DO - 10.1002/bit.10483

M3 - Article

C2 - 12491528

AN - SCOPUS:0037456181

VL - 81

SP - 430

EP - 437

JO - Biotechnology and Bioengineering

JF - Biotechnology and Bioengineering

SN - 0006-3592

IS - 4

ER -