Effects Of Low Temperature And Hormones On Glycerol-3-phosphate Acyltransferase(LpGPAT)gene Expression In Lilium Dauricum

Low temperature stress is a common disaster in flower cultivation,and it is one of the important factors affecting the quality of flower production.At the same time,hormones participate in the growth and development of flowers and the response mechanism of abiotic stresses such as low temperature,drought,and salt stress to provide resistance to flowers.The research on flower resistance is the need for scientific research to serve the production.Therefore,studying the resistance mechanism of flowers and grasping the law of gene expression changes under low temperature stress and hormone treatments have the advantages of improving flower production quality and breeding new resistant varieties Very important.Therefore,Lilium dauricum with strong cold tolerance and drought resistance is also an important wild resource for garden cross breeding.It is necessary to explore its specific resistance mechanism.Based on the early research of this research group,the Lp GPAT gene of Lilium dauricum was selected as the target gene for exploring the resistance mechanism of lily.In this study,phylogenetic tree construction was performed on the homologous GPAT amino acid sequences of 9 other species and the 10 GPAT amino acid sequences in Arabidopsis thaliana using bioinformatics analysis.Sequence alignment,analysis of the differences in the conserved domains of proteins,to explore the relationship between the structure and function of GPAT protein.The real-time fluorescence quantitative technology was used to analyze the transcriptional expression level of Lp GPAT gene in the leaves of Lilium dauricum under the induction of low temperature and different hormones(50 ?M abscisic acid,100 ?M salicylic acid,50 ?M ethephon,10 ?M methyl jasmonate).At the same time,according to the previous prediction results of cis-acting elements on the Lp GPAT gene promoter,transient expression vectors of four different promoter deletion fragments were constructed,and tobacco transformation was mediated by Agrobacterium,and GUS protein was subjected to low temperature and hormone stress.Qualitative and quantitative tests to determine whether the predicted cis-acting element can respond to low temperature and hormone signals in the roleof Lp GPAT gene transcription.Finally,the 1302-Lp GPAT-GFP subcellular localization vector was constructed for subcellular localization experiments to clarify the location of the Lp GPAT protein,to further understand the properties and functions of proteins,and the interaction between proteins is of great significance.The main results of this study are as follows:1.Lp GPAT protein and other plant GPAT proteins also have highly conserved acyltransferase active domains(Pls C): BOX I,BOX II,BOX III,and BOX IV,indicating that Lp GPAT protein is a member of the acyltransferase family and has acylase Of activity.The Lp GPAT protein and the Arabidopsis plastid GPAT(ATS1)protein are clustered into the same clade in the phylogenetic tree,so it can be speculated that Lp GPAT is localized on the plastids as ATS1.2.The Lp GPAT gene of Lilium grandis is rapidly expressed under low temperature stress and continues to be expressed under long-term low temperature treatment,which will lead to the metabolism of TAG in the leaves of Lilium dauricum and thus improve the low temperature resistance of Lilium dauricum.Under the induction of different hormones,Lp GPAT can respond to the induction of many plant hormones,which will help the lily to resist abiotic stress,which also provides an important theoretical reference for studying the function of Lp GPAT.3.There are many low-temperature and hormone-responsive cis-acting elements on the Lp GPAT gene promoter.According to the distribution position of the cis-acting elements,the promoter 5 'end deletion method was used to construct four promoter-deficient plant expression vectors(G1-G4).Agrobacterium tumefaciens-mediated transformation of Tobacco for transient expression analysis.Under normal conditions,the Lp GPAT promoter has a weak ability to start downstream GUS proteins,and G1> G2> G3> G4;under low temperature and induced by different hormones,the Lp GPAT promoter containing different cis-acting elements transcribes the GUS reporter gene There are differences in activity,but the ability to activate downstream GUS proteins is significantly stronger than normal conditions.This indicates that the Lp GPAT promoter is induced by low temperature and hormones,and Lp GPAT transcriptional activity is enhanced under abiotic stress to cope with adversity..4.The protein encoded by the Lp GPAT gene is located on the cell membrane and chloroplast.