PCR-identification of a Nicotiana plumbaginifolia cDNA homologous to the high-affinity nitrate transporters of the crnA family

Plant Mol Biol. 1997 May;34(2):265-74. doi: 10.1023/a:1005872816881.

Abstract

A family of high-affinity nitrate transporters has been identified in Aspergillus nidulans and Chlamydomonas reinhardtii, and recently homologues of this family have been cloned from a higher plant (barley). Based on six of the peptide sequences most strongly conserved between the barley and C. reinhardtii polypeptides, a set of degenerate primers was designed to permit amplification of the corresponding genes from other plant species. The utility of these primers was demonstrated by RT-PCR with cDNA made from poly(A)+ RNA from barley, C. reinhardtii and Nicotiana plumbaginifolia. A PCR fragment amplified from N. plumbaginifolia was used as probe to isolate a full-length cDNA clone which encodes a protein, NRT2;1Np, that is closely related to the previously isolated crnA homologue from barley. Genomic Southern blots indicated that there are only 1 or 2 members of the Nrt2 gene family in N. plumbaginifolia. Northern blotting showed that the Nrt2 transcripts are most strongly expressed in roots. The effects of external treatments with different N sources showed that the regulation of the Nrt2 gene(s) is very similar to that reported for nitrate reductase and nitrite reductase genes: their expression was strongly induced by nitrate but was repressed when reduced forms of N were supplied to the roots.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Anion Transport Proteins*
  • Bacterial Proteins / metabolism*
  • Base Sequence
  • Carrier Proteins / genetics*
  • Carrier Proteins / metabolism*
  • Chlamydomonas reinhardtii / genetics
  • Cloning, Molecular
  • DNA Primers
  • DNA, Complementary / isolation & purification*
  • DNA, Plant / isolation & purification
  • Fungal Proteins*
  • Gene Expression Regulation, Plant / drug effects
  • Genes, Plant*
  • Genome, Plant
  • Hordeum / genetics
  • Molecular Sequence Data
  • Multigene Family*
  • Nitrates / metabolism*
  • Nitrates / pharmacology
  • Nitrogen / pharmacology
  • Plant Roots / genetics
  • Plants, Toxic*
  • Polymerase Chain Reaction
  • Sequence Homology, Nucleic Acid*
  • Tobacco / genetics*

Substances

  • Anion Transport Proteins
  • Bacterial Proteins
  • Carrier Proteins
  • DNA Primers
  • DNA, Complementary
  • DNA, Plant
  • Fungal Proteins
  • Nitrates
  • nitrate transporters
  • Nitrogen

Associated data

  • GENBANK/Y08210