Molecular Cloning and Characterization of a Novel Gene Encoding DREB Protein from Buchloe dactyloides (Nutt.) Engelm

A 209bp expressed sequence tag (EST) was obtained with the degenerate primer technology, then based on the sequence of EST, the full-length cDNA of 1050 nucleotides was cloned from Buffalo grass by rapid amplification of cDNA ends (RACE). It was designated as BdDREB2, encoding a protein of 255 amino acids. The protein molecular weight was 28.39kDa, and the theoretical isoelectric point was 5.70. The BdDREB2 probably localized in nucleus and did not have signal peptide. A typical AP2/EREBP conserved domain was found in the coding region. The amino acid sequence compared by blast revealed high homology with DREB proteins of other plants, especially with the Buchloe-dactyloi DREB protein. The semi-quantitative analysis of expression of transcription showed: the expression of BdDREB2 gene reached the maximum after being treated by 20%PEG for 6h; BdDREB2 gene has the highest expression level in the root; the expression was significantly increased with the stress of drought (20% PEG) and high salt (3% NaCl), but not obvious with the stress of low temperature. Plant expression vector was constructed and transformed into the tobacco by the Agrobacterium-mediated methods. PCR amplification with the transgenic tobacco genome DNA indicated that the BdDREB2 gene had integrated in tobacco genome. RT-PCR showed that the BdDREB2 gene can be transcribed into mRNA in tobacco.