Transgenic Brassica carinata as a vehicle for the production of recombinant proteins in seeds

Hirudin, a blood anticoagulant protein from leeches, and beta-glucuronidase were produced in Brassica carinata Braun (Ethiopian mustard) seeds using oleosin as a carrier. Cotyledonary petioles were infected with Agrobacterium strains containing oleosin-glucuronidase (pCGNOBPGUS-A) or oleosin-hirudin...

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Bibliographic Details
Published in:Plant cell reports 1998-01, Vol.17 (3), p.195-200
Main Authors: Chaudhary, S, Parmenter, D.L, Moloney, M.M
Format: Article
Language:English
Subjects:
anticoagulants
beta-glucuronidase
Biological and medical sciences
biosynthesis
Biotechnology
Brassica carinata
cotyledons
drugs
Flotation
Fundamental and applied biological sciences. Psychology
gene expression
gene transfer
genes
Genetic engineering
Genetic technics
hirudin
Methods. Procedures. Technologies
Modification of gene expression level
petioles
protein synthesis
Proteins
recombinant proteins
Seeds
Transgenic animals and transgenic plants
Transgenic plants
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Summary:Hirudin, a blood anticoagulant protein from leeches, and beta-glucuronidase were produced in Brassica carinata Braun (Ethiopian mustard) seeds using oleosin as a carrier. Cotyledonary petioles were infected with Agrobacterium strains containing oleosin-glucuronidase (pCGNOBPGUS-A) or oleosin-hirudin (PCGN-OBHIRT) constructs. Polymerase chain reaction and neomycin phosphotransferase II enzyme assays confirmed the presence of the fusion genes in plants regenerating under selection. The fusion polypeptides were correctly expressed and targeted to the oil-bodies of the seeds with high fidelity (ca. 90%). Recombinant protein was purified from all other cellular protein by a simple flotation process and cleaved from oil-bodies using the endoprotease, Factor Xa. Hirudin activity was measured using a colorimetric thrombin inhibition assay and an activity in the range of 0.2-0.4 antithrombin units per milligram of oil-body protein was detected. B. carinata offers an attractive alternative for the production of recombinant proteins using oleosin technology.
ISSN:0721-7714
1432-203X
DOI:10.1007/s002990050377