Study On The Effect Of Plant Drought Tolerance Mechanism Of Soybean Phospholipase C Gene GmPLC10

Phospholipase C（PLC）is an important hydrolase indispensable to the metabolism and intracellular signal transduction both in plants and animals.It comprises three categories on the basis of substrate namely;Non-specific PLC（NPC）,phosphoinositide-specific PLC（PI-PLC）and glycosylphosphatidylinositol-specific phospholipase C（glycosylphosphatidylinositol PLC/GPI-PLC）.In plants,PLC lacks PH（Pleckstrin homology）domain making it similar to PLC-ζ1-5 in animals.Moreover,no corresponding receptors for DAG have been reported in plants to date.Instead,the content of IP₆ and the product of rapid phosphorylation of IP₃ were found in excess concluding that IP₆ and PA are the emerging second messengers in plants.PLC（GmPLC10）plays an important role in plant growth and development in normal as well as stressed conditions specifically drought.The recent reports about its role in drought resistance have attracted plant scientists.This study was aimed at demonstrating the key regulatory function of GmPLC10 gene in drought resistance.Agrobacterium tumefacians-mediated stable transformation in soybean cotyledons was performed using EHA105 strain containing the overexpression construct for GmPLC10 gene.The target gene was simultaneously transformed into Arabidopsis thaliana of which the phenotypic and physiological parameters were screened till T₃（Trangenic 3）generation.In the light of experimental and statistical analyses,we draw the following conclusions.1.We applied drought stress using 300 mM mannitol and compared the germination rate as well as primary root length in wt and transgenic Arabidopsis.The transgenic Arabidopsis showed quicker germination and longer primary root as compare to wt.2.Both the wild type and transgenic Arabidopsis were cultivated in soil for one week and the plants with same growth efficiency were selected and treated with drought stress for 10 days.Comparing the phenotypic and physiological parameters revealed depression in malondialdehyde content in transgenic Arabdidopsis while an increase in proline accumulation,slight increase in fresh weight and significant increase in dry weight and water content was observed.Na⁺level fell slightly in contrast to which K⁺level elevated while Na⁺/K⁺was reduced proving the key role of GmPLC10 in drought resistance.3.The degree of opening and closing of stomata in both wt and transgenic Arabidopsis were studied and compared.The stomatal aperture was larger in transgenic Arabidopsis than that of wt.The stomatal aperture of transgenic Arabidopsis were basically the same as that of wt with an additional PA and the stomatal aperture of wt were basically the same as that of transgenic Arabidopsis with an additional U73122 inhibitor.Hence,it is evident that GmPLC10 plays role in drought resistance.4.The Transcriptomic analysis of T₃ Arabisopsis plants showed differential regulation of 833 genes.Of 833 genes,281 were up-regulated while 452 were down-regulated.Through KEGG annotation enrichment analysis and GO analysis,we found 4 drought-related genes,but three of them was not found in the database thus demands further study.5.Using Agrobacterium-mediated genetic transformation of soybean,a total of 94T₀ resistant soybean plants were obtained,of which 12 plants were confirmed positive by PCR.At present,three T₁ generation positive plants have been obtained.