Functional Analysis Of MdCBF1 In Malus Domestica Borkh. CV. Fuji

Plants often endure lots of abiotic stresses in the group stages. such as drought, salinity, and low temperaturesand oxidative stress have enormous impact on agriculture and result in the deterioration of the environment. Plants have evolved to perceived different stress signals to transfer them and adapt to abiotic stresses by a variety of biochemical and physiological mechanisms, which to enhance the adaptability.Transcription factors play vital roles in sress signal transduction,gene expression modulation when plants are on the sress conditions.AP2/EREBPs are important transcription factors which related to plant development and physiology. It was investigated that AP2/EREBP transcription factors are able to improve tolerance to drought, high-salt and cold stresses in Arabidopsis and rice. CBF transcription factors belongs to AP2/EREBPs, which might be involved in low temperaturesand, drought, and high-salt abiotic stresses. But there is few report in Malus pumila Mill.Here, MdCBFI was isolated from CBF1 cDNA of Malus domestica Borkh.cv.Fuji which based on reported CBF1 cDNA. The results of entire nucleotide sequence analysis revealed that the full-length cDNA of MdCBF1 has a 660 bp open reading frame (ORF), encoding a predicted polypeptide of 219 amino acids with a molecular mass of 23.94kDa and calculated pI of 5.21. At the nucleotide sequence level, MdCBFl has higher sequence similarity. The protein contained an AP2 domain consisting of 58 amino acids. It contained the special aminophenol sequence of CBF protein with PKKRAGRKKFRETRHP in the 41th position and FADSAWR in the 113th position. MdCBFl also has the typical structure of CBF transcription factor. Two putative acidic domains in N-terminal and C-terminal regions were also detected. A putative nuclear localization signal located adjacent to C-terminal acid region.MdCBF1 was classified into DREB-A1 subgroup. The results of Real-Time PCR indicated that the expression of MdCBF1 gene was induced by drought stress and low-temperature, but was not by NNAand ABA. The transcript level of MdCBF1 gene was similar in Malus domestica Borkh.CV. roots, stems, leaves and seeds.We analyzed the function of expression of MdCBF1 gene in Arabidopsis thaliana plants, through constructing the Agrobacterium tumefaciens GV3101 containing plant binary expression vector. The floral dip method was used to transfer them into A. thaliana. The results showed that the MdCBF1 transgenic plants exhibited improved growth than wild type control plants on the low temperaturesand,but not evident on drought condition. The transgenic plants had higher proline. Moreover, they had lower electrolyte leakage percentage and lower malondialdehyde contents after cold stress. The expression level analysis of some stress-responsive genes indicated that some genes (e.g. RD29A and COR15a) were up-regulated.