Expression Characteristics Of The Proline Synthesis Key Gene P5CS1 In Poplar And Functional Verification Of Drought Tolerance

Poplars,as an important timber and afforestation species,are widely used in papermaking,furniture industry and bio-energy.The aboveground part of poplars grows fast and needs a number of water.However,the arid and semi-arid areas in northwest China lack of water,and the growth and development of poplars are severely inhibited.Drought has become an important factor affecting its sustainable development.Proline accumulation often occurs in response to osmotic stress,such as drought,to alleviate plant damage caused by stress.More and more researchs has been done on P5 CS,the gene that encodes a key enzyme for proline synthesis.In this study,we used bioinformatics,molecular biology and physiology techniques to clone the gene of proline synthesis,analyzed its spatiotemporal expression pattern and drought stress response,and identified the transgenic poplar lines at phenotypic,molecular and proline levels.The main results are as follows:1.The cloning analysis of P5CS1 gene of Populus alba × P.glandulosa showed that Pag P5CS1 was 2160 bp in length and composed of 719 amino acid codes.Bioinformatics analysis predicted that both Pt P5CS1 and Pt P5CS2 were non-transmembrane hydrophilic proteins without signal peptides in the existing database of Populus trichocarpa.Phylogenetic results showed that Pt P5CS1 and Pt P5CS2 proteins were highly conserved promoter elements in plant evolution,and Pt P5CS1 and Pt P5CS2 proteins contained a large number of cis-acting elements,indicating that P5 CS plays an important role in abiotic stress such as drought.2.The expression levels of poplar proline anabolism gene Pa P5CS1,Pa P5CS2 and Pa OAT were determined by fluorescence quantitative PCR.The results showed that the expression levels of poplar proline anabolism gene P5CS1 gradually increased with the increase of drought extent.P5CS1、P5CS2 and OAT were both expressed in the roots,stems and leaves of poplar,in which P5CS1 and P5CS2 had the highest expression level in leaves and the lowest expression level in stems.OAT expression level was highest in poplar leaves and lowest in roots.3.The Pag P5CS1 overexpression vector was constructed,and 12 poplar lines were successfully obtained by agrobacterium mediated transformation of poplar leaves.The highest expression of P5CS1 gene was 8.3 times that of the wild type.4.Compared with the wild type,leaf wilting degree of overexpressed lines was lower during drought stress treatment.Under drought stress,the expression level of P5CS1 gene and proline content in the overexpressed lines were higher than those in the wild type,indicating that the overexpression of Pag P5CS1 enhanced the drought tolerance of poplar,and P5CS1 played a positive regulatory role under drought stress.In conclusion,P5CS1 gene is highly responsive to drought stress,and poplar with Pag P5CS1 overexpression can enhance drought tolerance by increasing proline accumulation.