Abstract

Nitrate reductase is a key enzyme in the overall process of nitrate assimilation by plants. A nitrate reductase (NR) gene related to somatic embryogenesis was firstly cloned from Gossypium hirsutum. Sequence alignment shows that the GhNR is closest with Tilia platyphyllos NR gene. The expression of GhNR was investigated using semi-quantitative RT-PCR in nonembryogenic calli, embryogenic calli and somatic embryoids which showed that the expression level of GhNR was higher in embryogenic calli and somatic embryoids than in nonembryogenic calli. Besides, the expression of GhNR gene was verified by transformation in E. coli BL21 (DE3) strain with the recombinant expression vector pET-32a. NR activity assay showed that the crude GhNR protein had obvious activity to KNO₃ substrate. The results implied that NR has a critical function in the assimilation of nitrogen in cotton somatic embryo germination.

Key words: Cotton, somatic embryogenesis, nitrate reductase, gene expression.

Abbreviation

RT-PCR, reverse transcriptase-PCR; NR, nitrate reductase;GhNR, nitrate reductase gene cloned from Gossypium hirsutum; ORF, open reading frame; IPTG, isopropylthiogalactoside; SDS-PAGE, sodium dodesyl sulfate-polyacrylamide gel electrophoresis.