The-Soybean Proline Synthase GmP5CS Gene-Involved In The Preliminary Functional Study Of Salt Tolerance

As an important cash crop in the world,soybean(Glycine max)plays an important role in human life.With the increasing of soil area,the adaptability of soybean to salt stress needs to be strengthened.Proline maintains plant’s normal metabolism by a large amount of accumulation on salt stress which depends on P5 CS regulating the whole proline synthesis pathway.In order to further understand the molecular mechanism of the involvement of Gm P5 CS gene in salt tolerance,the Gm P5 CS full-length vector was constructed,accompanying with relative gene expression,transformation of Gm P5 CS transgenic "composite" plants and function researches such as observation of phenotype,calculation of survival rate,determination of relative water content,proline content and antioxidant enzyme activity of transgenic "composite" plants were determined under salt stress and semi-quantitative western-blot.The results show below:(1)Real-time quantitative PCR was used to analyze the expression pattern of Gm P5 CS gene under salt stress,the results showed that the relative expression of Gm P5 CS was significantly upregulated in response to salt stress,and the relative expression of Gm P5 CS at three hours and six hours was significantly higher than the control,which showed that the Gm P5 CS gene was triggered by its own promoter and enhanced the salt tolerance of transgenic "composite" plants.(2)The Gm P5 CS full-length vector was constructed,and the Gm P5 CS transgenic "composite" plants were obtained by Agrobacterium rhizogenes-mediated hairy roots transformation and cotyledon node transformation,and the positive transgenic hairy roots were identified and screened by confocal microscopy.There was no difference between the control and the Gm P5 CS transgenic hairy roots under bright light,but the Gm P5 CS transgenic hairy roots had green fluorescence under the blue light excitation of confocal microscope,finally,the positive transgenic hairy roots were obtained.(3)The phenotype,survival rate and relative water content of Gm P5 CS transgenic "composite" plants were observed,calculated and determined after six days of treatment with 200 mM NaCl solution.The results showed that the Gm P5 CS transgenic "composite" plants showed healthier leaves and longer roots than the control,and significantly increased the survival rate and relative water content of the Gm P5 CS transgenic "composite" plants.(4)After being treated with 150 mM NaCl solution for six hours,the related physiological indexes were determined,the results showed that the proline content,catalase,peroxidase and superoxide dismutase activity of Gm P5 CS transgenic "composite" plants were significantly higher than the control,which showed that the Gm P5 CS transgenic "composite" plants responded to salt stress by increasing proline content,peroxidase,catalase and superoxide dismutase activity.(5)After being treated with 150 mM NaCl solution for six hours,the expression of Gm P5 CS protein in transgenic hairy roots was analyzed by semi-quantitative western blot,which showed that salt stress increased the expression of Gm P5 CS protein.