Nucleotide sequence of a spinach nitrate reductase cDNA

The assimilation of nitrate is energetically expensive and the expression and activity of the enzymes in the pathway are tightly regulated. Nitrate reductase (NR: E.C.1.6.6.1.) is the first enzyme in this pathway, catalysing the rate-limiting step and is considered to be the key site of regulation....

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Bibliographic Details
Published in:Plant molecular biology 1990-07, Vol.15 (1), p.187-190
Main Authors: Prosser, I.M. (Bristol Univ. (UK). Dept. of Agricultural Sciences), Lazarus, C.M
Format: Article
Language:English
Subjects:
ADN
Amino Acid Sequence
ANTICORPS MONOCLONAL
ANTICUERPOS MONOCLONALES
ARN
Base Sequence
Biological and medical sciences
Cloning, Molecular
DNA
DNA - genetics
Fundamental and applied biological sciences. Psychology
Genes. Genome
Molecular and cellular biology
Molecular genetics
Molecular Sequence Data
MONOCLONAL ANTIBODIES
NITRATE REDUCTASE
Nitrate Reductases - genetics
NITRATO REDUCTASA
NUCLEOTIDE SEQUENCE
Plants - enzymology
Plants - genetics
RNA
SECUENCIA NUCLEICA
SEQUENCE NUCLEIQUE
SPINACIA OLERACEA
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Summary:The assimilation of nitrate is energetically expensive and the expression and activity of the enzymes in the pathway are tightly regulated. Nitrate reductase (NR: E.C.1.6.6.1.) is the first enzyme in this pathway, catalysing the rate-limiting step and is considered to be the key site of regulation. NR is induced by its substrate, nitrate. We set out to clone spinach NR. Total RNA was extracted from leaves of nitrate-fed plants and mRNA isolated by oligo-dT-cellulose chromatography. mRNA from the peak fraction was used to prepare cDNA, which was cloned in lambda gt11 via Eco RI adapters. The library was screened with the squash NR cDNA and two clones with inserts greater than 1.8 kb were identified and subcloned into pUBS1 (modified Bluescript). DNA sequencing and comparison with published data have confirmed that both clones (pIMP4, insert = 3.3 kb and pIMP5, insert = 1.9 kb) are derived from NR mRNAs. pIMP4 encodes an open reading frame of 926 amino acids (104 kDa) with 79, 75 and 67% identity (88, 87, 81% similarity) to tobacco, Arabidopsis and rice respectively.
ISSN:0167-4412
1573-5028
DOI:10.1007/BF00017743