Isolation And Characterization Of Trehalose-6-Phosphate Phosphatase Gene From Caragana Korshinskii

Caragana korshinskii Kom. is mainly distributed in the western region of Inner Mongolia, which is a sand-fixied and soil and water-conserved plant as well as fodder shrub in the desert and desert steppe zone, and shows widely adaptable and strong stress-tolerant. Trehalose is a non-reducing disaccharide that plays an important role in stress-tolerance of plants. Under the abiotic stress conditions, trehalose as an osmotic substance keeps plant from damage. Trehalose-6-phosphate phosphatase (TPP) is the key enzyme in trehalose metabolism pathway, which catalyzes the dephosphorylation of trehalose-6-phosphate phosphatase to generate trehalose. The expression of TPP directly affects trehalose synthesis, therefore it has great practical significant to study function and characterization of CkTpp genes for genetic improvement of crops and forest trees.In this study, CkTpp cDNA was isolated from Caragana korshinskii Kom. by homology cloning technique. The full-length of CkTpp cDNA contains 1004bp, including an open reading frame of 855bp, encodes a protein of 284 amino acids with a molecular weight of 32.3KdDa, and a theoretical isoelectric point of 9.2. Phylogenetic analysis showed that CkTpp is in a sister group to the clade of Tpps from Leguminosae sp. such as Glycine max, Glycine soja and Cicer arietinum. To investigate the function and expression characteristic of the CkTpp, prokaryotic expression vector pET-32a(+)-CkTpp was constructed, and abiotic stress experiments were performed with the prokaryotic expression strain. The results showed that tolerance of transgenic strain overexpressing CkTpp to abiotic stress, such as drought, thermal and salt were enhanced. On the other hand, binary expression vector pBI101-CkTpp was constructed by homologous recombination, which provided a foundation for analyzing function of the CkTpp by transforming Arabidopsis further.