| Salinization,drought and other environmental factors seriously affect crop growth and yield.And they all affect the water retention capacity,lead the crop to death.Along with the population unceasing increase,the cultivated area daily reduction and the ecological environment suddenly worsening,the grain supply and demand will be contradictory day by day.How therefore enhances the crops resist drought,high salt and high alkali ability increases the grain yield is the question which one urgently awaits to solve.Compared with traditional breeding techniques and molecular marker-assisted breeding,it is more promising to introduce plant species to improve plant resistance by genetic engineering.On the basis of the preliminary research in this laboratory,soybean GmMMK2,GmPP1 and GmMLP were found to be regulated by salt stress at the protein level by i TRAQ technique.GmMMK2 is MAPK kinase protein,GmPP1 is Ser/Thr protein phosphatase,GmMLP is belong to Bet ? protein,a Major latex protein,The function of the three target genes under abiotic stress is still unknown.Since GmMMK2,GmPP1 and GmMLP were induced by salt stress at the protein level,we speculated that the three target genes could also perform corresponding biological functions in other stress signaling transduction processes.Aiming at those problems,this study constructed plant expression vectors of GmMMK2,GmPP1 and GmMLP.And the tobacco was transformed by Agrobacterium Tumefaciens.There are many transgene plants were obtained.The biological function of transgenic tobacco T1 generation plants was studied,and the function of genes under drought and high salt stress was analyzed.To study the response mechanism of GmMMK2,GmPP1 and GmMLP genes under drought and high salt stress and to cultivate new strains of resistant genetically modified crops,the genetic basis was provided and the gene material was provided.The main results was summarized as follows:(1)The GmMMK2,GmPP1 and GmMLP genes were successfully cloned in soybean,And the expression patterns of three target genes under salt,alkali and ABA were analyzed.The study found that the three target genes had different responses to the three stress treatments.The expression patterns of GmMMK2,GmPP1 and GmMLP genes in different soybean tissues were also analyzed.(2)There were three plant expression vectors were constructed,they were pCAMBIA2300-GmMMK2,p CAMBIA2300-GmPP1 and p CAMBIA-GmMLP.The plant expression vectors of the constructed three genes were transferred into tobacco by Agrobacteriumtumefaciens-mediated transformation,respectively.There were 34 PCR positive and 29 RT-PCR positive plants be obtained,which were transformed with p CAMBIA2300-GmMMK2;there were14 PCR positive and 13 RT-PCR positiv plants be obtained,which were transformed with pCAMBIA2300-GmPP1;there were 35 PCR and 23 RT-PCR positivepositive plants be obtained,which were transformed with p CAMBIA-GmMLP.Km screening and molecular biology detection of transgenic T1 lines.Five transgenic lines of GmMMK2 gene was detected by PCR,three transgenic lines of GmPP1 gene was detected by PCR,six transgenic lines of GmMLP gene was detected by PCR.(3)The transgenic T1 generation lines were subjected to abiotic stress treatment,The results showed that the transgenic tobacco plants of GmPP1 and GmMLP genes increased the root length of tobacco at seedling stage compared with non-transgenic tobacco plants under salt stress treatment.Under the 200 m M mannitol treatment,the resistance to osmotic stress ability of expression of GmMLP gene was stronger than that of wild type plant.Under drought stress,the tobacco with expression of GmMLP gene increased tolerance to drought at seedling stage compared with non-transgenic tobacco.Suggesting that GmPP1 gene and GmMLP gene have the function of improving plant salt tolerance and drought resistance respectively. |
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