Cloning And Expression Analysis Of PhRAX2 And PhRAX3 In Petunia

Petunia hybrida belong to solanum of solanaceae,with rich color and long flowering period,is widely applied in gardens.At the same time,petunias are easy to be transgenic and have a short growth cycle.Therefore,it has become a model plant for shoot branching and stress tolerance research.In the cultivation process of petunia,multiple topping is needed to control the plant type,which greatly increases the production cost.Therefore,developing new varieties with multiple branches is the key to efficient production of petunia.In addition,drought is the main factor limiting the application range of petunia.Therefore,it is an important research direction of petunia to cultivate new varieties with strong stress resistance.Iii order to preliminarily explore the functions of PhRAX2 and PhRAX3 in petunia,the full-length sequences of PhRAX2 and PhRAX3t as well as the promoter sequence of PhRAX3 were cloned,their bioinformatics were analyzed,and the expression characteristics of PhRAX2 and PhRAX3 in different tissues and different treatments were studied.The PhRAX2,PhRAX3 overexpression vector and pPhRAX3 fusion GUS vector were constructed and transferred into agrobacterium.The results are as follows:1.The R2R3-MYB family gene PhRAX2 was cloned.The total length of the gene was 864 bp,and the similarity with NtRAX2 of tobacco was as high as 70%.The 1R-MYB family gene PHRAX3 was cloned,with a total length of 1539 bp,which was highly similar to the first 80 amino acid sequences of tomato S1RAX3,pepper CaRAX3 and other species.The promoter sequence of PhRAX3 was cloned,with a total length of 1917 bp.The sequence contains several regulatory elements of drought response and auxin response.2.Tissue specific expression analysis of PhRAX2 and PhRAX3:(1)the expression level of PhRAX2 in apex was highest,which was about 2 times of the roots,the expression level in leaf axils and leaves was about 1.3 times of the roots,and the expression in stems was about 1/2 of the roots.(2)the highest expression level of PHRAX3 was found in roots,and the expression levels in leaves,stems,stem tips and leaf axils were 2/5,1/5 and 1/10 of roots,respectively.3.Analysis of the regulation of PhRAX2 by exogenous cytokinins:after 6-BA treatment for 6 h,the expression of PhRAX2 increased to 1.2 times that of the control and 3.5 times that of the control after 12 h,indicating that PhRAX2 can be positively regulated by cytokinin.4.Analysis on the regulation of PhRAX3 by simulated drought:after PEG treatment for 6 h,the expression of PhRAX3 increased to 12 times that of the control,increased to 38 times that of the control after 12 h,and decreased to 1.9 times that of the control after 24 h,indicating that PhRAX3 can respond to drought stress treatment rapidly.5.PhRAX2 and PhRAX3 were respectively connected to the PJL-blue vector,which was then linked to PFK209 after LR reaction,and the overexpression vector of 35S::PhRAX2 and 355::PhRAX3 was constructed.The promoter sequence of PhRAX3 was connected to the pCambia1391Z vector to construct the pPhRAX3:GUS vector.Three vector plasmids were transferred into agrobacterium and transformed into arabidopsis wild-type,which laid a foundation for subsequent experiments.