Molecular cloning, characterization, expression and chromosomal location of OsGAPDH, a submergence responsive gene in rice (Oryza sativa L.)

Abstract.

Differential clones from submergence stress and control treatment from rice seedlings were isolated by the differential screening method. One of the clones, OsGAPDH, represented a gene that was expressed at high level during 12-h submergence. A homology search of GenBank databases showed that OsGAPDH had significant sequence homology with maize non-reversible glyceraldehyde-3-Phosphate dehydrogenase. The OsGAPDH sequence consists of 1,772 bp with the longest open reading frame encoding 499 amino acids with a calculated relative mass of 54.2 kDa. Genomic Southern analysis indicated that one or two copies of the OsGAPDH gene occur in the Yukihikari genome. The chromosomal location of the OsGAPDH gene was identified by RFLP analysis indicating that OsGAPDH was located on chromosome 8. Tissue-specific expression of OsGAPDH indicated that the high level of mRNA was detected in the panicle. Plants exposed to drought, submergence and ABA treatment showed an increased accumulation of OsGAPDH transcripts. The induction of Escherichia coli cells containing the pGST-OsGAPDH plasmid resulted in the accumulation of a large amount of the 83.2-kDa recombinant protein. The purified GAPDH enzyme showed an optimum activity at pH 8.5 and 50 °C, and was strongly inhibited by ATP and ADP.