| Land area affected by salinization in the world has reached 10%.Salt stress is the second only to drought caused agricultural yield loss,leading to serious loss of modern agricultural production.Application of genetic engineering to improve varieties,thus improving salt tolerance of the plant is a rapid and effective method.Transferring salt-tolerant genes into plants,salt tolerance of glycophyte might be improved,references for cultivating new varieties of salt tolerant plants and making full use of saline soil were also provided.CFLAP1 is a member of bHLH transcription factor family,in the ABA-dependent signalling pathway under hyperosmotic stress,ABA and PYR/PYLs promote the activation of downstream PP2 C phosphokinase,PP2 C then activates SnRK2,SnRK2 could phosphorylate CFLAP1,CFLAP1 activates downstream transcription factor to adapt to stress pressure and so on.In our current research,the CFLAP1 gene of Arabidopsis thaliana was cloned and its role in salt tolerance was verified.A 2×35S-CFLAP1-GFP-Nos overexpression vector was constructed by Gateway technique,Arabidopsis thaliana transformed by floral dipping method.T1,T2 and T3 transgenic lines were screened by hygromycin(37.5 ?g/ml)resistance,and finally the overexpression CFLAP1 homozygous transgenic Arabidopsis thaliana plants were obtained.DNA expression level revealed that the fusion gene was successfully integrated into the genome of Arabidopsis thaliana.Western blot indicated that fusion protein GFP-CFLAP1 was stably expressed.Fluorescence microscopy showed that the fusion protein was located on the Arabidopsis root cell membrane.Branch numbers of overexpress CFLAP1 transgenic Arabidopsis were more than wild type via phenotype observation.Following salt treatment of transgenic and wild-type Arabidopsis thaliana seedlings(salt concentrations were 0 mM,100 mM,150 mM and200 mM,respectively),physiological indicators were determined.The results were as follows:(1)With the increase of salt concentration,the fresh and dry weight of both transgenic Arabidopsis thaliana and control were decreased,but transgenic Arabidopsis thalianawere higher than those of control group at same salt concentration.(2)Root length of transgenic Arabidopsis thaliana was longer than that of the control group under 4corresponding salt concentrations and was significantly different at concentrations of150 m M and 200 mM: transgenic Arabidopsis thaliana increased 34 % and 30 % at salt concentrations of 150 m M and 200 mM,compared to control,respectively.(3)The contents of proline in Arabidopsis thaliana and control group were increased with the increase of salt concentration,and the content of proline in Arabidopsis thaliana was 28.10 % and 68.70% higher than that in control group at the concentrations of 150 m M and 200 m M : the content of proline were 22.40 ?g/g and28.70 ?g/g at salt concentration of 150 mM in transgenic and control,respectively;and the content of proline were 89.19 ?g/g and 52.84 ?g/g at salt concentration of 200 mM in transgenic and control,respectively.(4)The contents of malonaldehyde in Arabidopsis thaliana and control group were increased with the increase of salt concentration,while the content of MDA in Arabidopsis thaliana was 21.30 % and33.08 % lower than that in control at the salt concentrations of 150 mM and 200mM:contents of MDA in the transgenic and control groups were 16.89 nmol/g and21.47 nmol/g at the salt concentration of 150 mM,respectively,and the content of MDA in the transgenic and control groups were 23.85 nmol/g and 35.65 nmol/g at the salt concentration of 200 mM,respectively.The root length,dry weight and fresh weight of overexpression CFLAP1 transgenic Arabidopsis were all higher than that of wild type under salt stress,and the content of proline and MDA showed the salt tolerance of over-expression CFLAP1 transgenic Arabidopsis thaliana was higher than that of wild type.All the data suggested that overexpression of CFLAP1 could enhance salt tolerance of Arabidopsis thaliana. |
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