PCR-identification of a Nicotiana plumbaginifolia cDNA homologous to the high-affinity nitrate transporters of the crnA family.
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.
Quesada
A
A
I.N.R.A., Laboratoire de Biologie Cellulaire, Versailles, France.
Krapp
A
A
Trueman
L J
LJ
Daniel-Vedele
F
F
Fernández
E
E
Forde
B G
BG
Caboche
M
M
D000595:Amino Acid Sequence; D000818:Animals; D027321:Anion Transport Proteins; D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism*; D016825:Chlamydomonas reinhardtii / Q000235:genetics; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary / Q000302:isolation & purification*; D018744:DNA, Plant / Q000302:isolation & purification; D005656:Fungal Proteins; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D017343:Genes, Plant; D018745:Genome, Plant; D001467:Hordeum / Q000235:genetics; D008969:Molecular Sequence Data; D005810:Multigene Family; D000091285:Nitrate Transporters; D009566:Nitrates / Q000378:metabolism* / Q000494:pharmacology; D009584:Nitrogen / Q000494:pharmacology; D018517:Plant Roots / Q000235:genetics; D010947:Plants, Toxic; D016133:Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid; D014026:Nicotiana / Q000235:genetics*
D027321:Anion Transport Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; D017931:DNA Primers; D018076:DNA, Complementary; D018744:DNA, Plant; D005656:Fungal Proteins; D000091285:Nitrate Transporters; D009566:Nitrates; D009584:Nitrogen