Abstract:

A cucumber cDNA designated CsNMAPK (GenBank accession NO. DQ812086) was isolated by RT-PCR and RACE. Bioinformatics analysis indicated that the full-length cDNA sequence was 1 636 bp, which contained an open reading frame (ORF) of 1 113 bp and encoded a protein of 370 amino acid residues. The subcellular localization analysis predicted that the protein was in the cytoplasm and there was one strong inside to outside transmembrane helix. PlantCare analysis indicated that there were abscisic acid, MeJA, salicylic acid cis-acting responsive elements and gibberellin, wound-responsive elements in CsNMAPK. The CsNMAPK was cloned into pET-30a(+) vector to construct recombination prokaryotic expression vector pET-CsNMAPK. After transformed to E.coli BL21 and induced by isopropyl-β-D-thiogalactopyranoside (IPTG), recombinant protein about 46 kD was expressed in pET-CsNMAPK system and separated by SDS-PAGE electrophoresis. The research will be helpful to study the function of cucumber MAPK gene in response to NO3- stress.

Key words: cucumber, MAPK, NO₃^-stress, bioinformatics analysis, prokaryotic expression