Drought-resistant Gene Expression And Proteome Analysis Of Potato Under Drought Stress

Potato(Solanum tuberosum L.)is currently planted by more than 150 countries as the fourth staple crops in the whole world.In China,both the potato planting area and yields rank number one in the world,and most of the potato are planted in the drought and semi-drought area.However,the potato is very sensitive to the water shortage.With the gradual deterioration of the environments and the water shortage,the industry of potato is threatened severely.It is the first selection of high and stable potato yield that we Breed drought-resistant potato varieties and improve thier drought tolerance.In this study,Virus free test-tube Plantlets of Jinshu 2(with the strongest drought resistance),Dongnong 308(with the medium drought resistance),and Favorita(with the worst drought resistance)with the treatment of 20%PEG-6000 to mimic drought stress were applied to elucidate the effects of drought on the ultrastructure and physiological parameters of potato,which is helpful to evaluate the extents of drought resistance.STAREB gene,regulatory transcription factor was cloned by RT-PCR,and its expression under drought conditions was studied;the proteomics of potato leaves under drought condition was studied-by 2D SDS-PAGE;the expression patterns of drought response genes were detected by real-time quantative PCR.In all,the whole study is trying to elucidate the drought resistance mechanisms,and to provide theoretical evidences for the studies of crop drought resistance and breeding.The results are as follows:1.The chloroplast was the most sensitive to drought and is impaired the most,while the mitochondria was more stable than the chloroplast;the damage of the ultrastructure of stem cells was less than that of the mesophyll cells;the damage of the ultrastructurc of drought resistant varieties was less than that of the drought sensitive varieties.The various physiological parameters changed with the alteration of ultrastructure.Under drought condition,the amounts of MDA increased by 48.89%?243.27%,and the differences are significant in all 3 varieties;the activities of POD increased by 2.06%?32.98%,and the differences were very significant and significant in Favorita and Jinshu 2,respectively;The relative water content and the activities of root decreased;the alteration of SOD and chlorophyll were different among 3 varieties.In summary,the content of MDA and the damaged chloroplast ultrastructure of plantlets in vitro were changed significantly,and these changes can be well correspond to the levels of drought resistance of these tested varieties.Damage degree of chloroplast and content changes of MDA can be used as a potato drought resistance material screening effective index.2.StAREB gene was cloned by RT-PCR method,and the blast showed that it shares 99.71%similarity with the other StAREB gene found in NCBI.There are 3 conserved phosphorylation sites(C1,C2,and C3),a nuclear targeting signal NLS(KVVE)and a conserved bZIP domain in the encoded StAREB amino acids.The 3 conserved phosphorylation sites share high similarities with them showing in the AtABFl,AtABF2,AtABF3,AtABF4,NbZIP,HvABI5,and OsABI5.StAREB is the most close to.SIAREB evolutionarily,following by AtABF2 and NtbZIP,but not very close with the other bZIP transcription factors,such as HvABI5,OsABI5,AtABF1,AtABF3,and AtABF4.StAREB gene was induced by drought(PEG-6000),low temperature,and high salt.Under drought condition,StAREB gene was strongly induced,and the expression levels stayed at very high levels during the whole drought treatment.3.After 2D SDS-PAGE,16 different expressed spots of proteins were acquired.Among these 16 proteins,proteins of number 1-14 were upregulated,including phosphorylation kinase,haloacid dehalogenase,heat shock protein 70 kDa,heat shock protein STI,the ?subunit of ATP synthase,soluble inorganic pyrophosphatase 1,glycine rich protein,ascorbate dehydrogenase,ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit,chloroplast manganese stabilizing protein,glyceraldehyde-3-phosphated ehydrogenase B,50S ribosomal protein L4 protein,Chain B,D of Ribulose-5-Phosphate 3-Epimerase.In contrast,proteins of number 15 and 16 were downregulated,including isocitrate dehydrogenase[NAD+]regulatory subunit 1 and disulfide-isomerase precursor-like protein.After analyzing the proteins found in this study,the proteins were classified into 5 groups,including photosynthesis,the maintenance and regulation of energy metabolism,the rapid response to drought stress,and the elimination of ROS and the enhancement of defense.4.11 genes were consistent with the expression of their derived proteins,which 10 genes were upregulated and one was downregulated after 48 h drought treatment.while 5 genes expressed inconsistently with their encoding proteins.Among them,the genes encoding haloacid dehalogenase,heat shock protein 70 kDa,chloroplast manganese stabilizing protein,and glyceraldehyde-3-phosphatede hydrogenase B were downregulated versus the upregulation of their derived proteins.In contrary,the isocitrate dehydrogenase[NAD+]regulatory subunit 1 was upregulated with the downregulation of its encoding protein.The genes expressed very differently in the testing varieties.Some of the genes were induced in both varieties;some of the other genes were highly induced in the drought resistant variety,while not induced or highly inhibited in drought sensitive variety.