Cloning And Stress Resistance Analysis Of CBF2 Gene From Almond

Genus of subgenus Amygdalus almond,Rosaceae,peach almond also known as Li Yake,is the world famous fruit oil trees.Almond is a perennial fruit trees,the growing season especially in flowering period.Once it is susceptible to low temperature freezing,chilling injury,often occur in flower bud withered and dry branches,affect the normal growth of trees and strong,causing serious economic losses.In this study,a gene encoding the CBF transcription factor was isolated from almond with PCR based on bioinformation databases.named AcCBF2.The main results of this study are as follows:1 Sequence analysis showed that the full-length of AcCB2 was a 717 bp open reading frame which encoded a protein of 238 amino acids with predicted molecular weight of 26.59 kD and a isoelectric point of 5.29.Multiple sequence analysis showed that AcCBF2 protein share high similarity with those plant cold stress-related CBF proteins in other plants,which contain the classic AP2 domain and two CBF signature sequence.Phylogenetic analysis revealed that the isolated almond AcCBF2 gene can be classified into A-1 subgroup of the CBF subfamily.2 The recombinant prokaryotic expression vector Pet-AcCBF2 was constructed by inserting the DNA fragment into the prokaryotic expression vector Pet-32a(+),and then transformed into E.coli Rosetta(DE3).Prokaryotic expression of AcCBF2 showed that AcCBF2 could be expressed in prokaryotic cells.3 The AcCBF2 gene was inserted into the yeast expression vector of pGBKT7,The results showed that AcCBF2 had the activation activity in yeast cells.4 RT-PCR analysis indicated that the expression of AcCBF2 gene only have response to low temperature and drought,no response to salt stress and ABA.5 The plant expression vector p1304-AcCBF2 was transformed into Arabidopsis using floral dipping method.T3 generation transgenic Arabidopsis was used to analyze the stress characteristics The results indicated that overexpression of AcCBF2 enhanced tolerance to low temperature and drought in transgenic Arabidopsis,but AcCBF2 was no response to high salt stress.