Cloning and characterization of a cDNA encoding aspartate aminotransferase-P1 from Lupinus angustifolous root tips

A root tip cDNA library, constructed in the lambda Zap II expression vector, was immunoscreened with a monoclonal antibody raised against aspartate aminotransferase-P1 from Lupinus angustifolius L. var Uniharvest. One 1452-base pair clone was isolated. The encoded protein sequence had high homology...

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Bibliographic Details
Published in:Plant physiology (Bethesda) 1994, Vol.104 (2), p.417-423
Main Authors: Winefield, C.S, Reddington, B.D, Jones, W.T, Reynolds, P.H.S, Farnden, K.J.F
Format: Article
Language:English
Subjects:
amino acid sequences
aspartate transaminase
cloning
cytosol
DNA
Escherichia coli
genbank/m92094
gene expression
gene transfer
genetic code
genetic techniques and protocols
genetic transformation
Lupinus angustifolius
molecular sequence data
mutants
nucleotide sequences
root tips
Saccharomyces cerevisiae
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Summary:A root tip cDNA library, constructed in the lambda Zap II expression vector, was immunoscreened with a monoclonal antibody raised against aspartate aminotransferase-P1 from Lupinus angustifolius L. var Uniharvest. One 1452-base pair clone was isolated. The encoded protein sequence had high homology to both plant and animal aspartate aminotransferase sequences. The clone was converted to the phagemid form and expressed in Escherichia coli. The expressed protein was enzymically active and could be immunocomplexed with aspartate aminotransferase-P1-specific antibodies. The complete aspartate aminotransferase-P1 cDNA was cloned into the yeast expression vector pEMBL-yex4 and transformed into Saccharomyces cerevisiae strain BRSCS6, which possesses a mutated aspartate aminotransferase gene (the asp5 mutation). Complementation of the mutation was achieved using this construct.
ISSN:0032-0889
1532-2548