Function Analysis Of PuAHL17 Gene For Drought Tolerance In Populus Ussuriensis

At-hook protein plays a crucial role in plant growth and development,organ construction,abiotic stress,hormone signaling and other responses.AT the same time,AT-Hook protein can also regulate transcription activities of transcription factors or assist transcription factors to improve their transcription activities.Studies have reported that AT-hook proteins,as transcription factors,can antagonize with photochromin interaction factors(PIFs)to promote the transcription of growth and hormone-related genes.In addition,in previous reports,most of the studies on AT-hook proteins were conducted on herbaceous plants,while there were relatively few studies on woody plants.Therefore,in this study,the protein prediction of Populus ussuriensis was analyzed by bioinformatics method.At the same time,the expression pattern of PuAHL17 gene in Populus ussuriensis was elucidated under drought stress,and its drought-resistant function was studied.The main results are as follows:(1)The expression pattern of PuAHL17 gene was analyzed by real-time quantitative fluorescent PCR(q RT-PCR)under the stress of 6% PEG6000,abscisic acid(ABA),methyl jasmonate(MeJA),salicylic acid(SA)and sodium chloride(Na Cl).The results showed that under drought stress simulated by 6% PEG6000,the expression of PuAHL17 gene was significantly up-regulated in both the roots and leaves of Populus ussuriensis,and the expression level in the leaves was higher than that in the roots.The function of PuAHL17 gene promoter was predicted by Plant Care online analysis,and the promoter region was found to contain ABRE,ARE,MRE and MBS,which are related to drought stress response.The promoter sequence of PuAHL17 gene was cloned from the genome of Populus ussuriensis.pro PuAHL17::GUS transgenic plants were obtained by leaf disk method mediated by Agrobacterium tumefaciens,and GUS staining was performed on the transgenic plants.The results showed that the roots and leaves of Populus ussuriensis had light staining without stress.After drought stress simulated by 6% PEG6000,the staining in roots and leaves was significantly deepened.(2)Subcellular localization experiment showed that PuAHL17 gene transcription factor of Populus ussuriensis was localized in the nucleus.The transcriptional activity of PuAHL17-BD vector was analyzed using the transient expression method of the protoplast of Populus alba.PuAHL17 showed no transcriptional activity.(3)Construct 35 S promoter-driven PuAHL17 overexpression(PuAHL17-OE)vector and35 S promoter-driven PuAHL17 inhibitory expression(PuAHL17-RNAi)vector,and use agrobacterium tumefaciens mediated leaf disk method for genetic transformation of Populus ussuriensis.A total of 14 PuAHL17-OE lines and 4 PuAHL17-RNAi lines were obtained.The drought resistance of WT and PuAHL17-OE strains was analyzed and compared under 6%PEG6000 stress.The results showed that under 6% PEG6000 simulated drought stress,the dry weight of root and leaf of PuAHL17-OE was heavier than that of WT,the contents of pconductivity,hydrogen peroxide and malondialdehyde in root,stem and leaf were significantly decreased,while the contents of proline,peroxidase and superoxide dismutase were significantly increased.In conclusion,PuAHL17 is a transcription factor localized in the nucleus,and its overexpression can improve drought resistance of Populus ussuriensis by increasing the clearance capacity of reactive oxygen species(ROS).