Study On Differential Expression Genes Of Phytophthora Sojae During Sporangium Formation Process

This study used Phytophthora sojae isolates using cellophane and carrot agar medium to grow abundant mycelium,and analyzed to the profile of differential genes expressed in early stage(0-7h) of sporangium formation under sterilized water condition by using mRNA differential display reverse transcription PCR.The study was identified to the effect of regulation genes expressed in sporulation process though the reverse northern hybridization,cloning,analysis of sequence, comparing homology and functional analysis of these differential fragments.The study established basis for controlling of sporulation of P.sojae.The results showed that;The growth of P.sojae using cellophane and carrot agar can obtained pure vegetative mycelium;the pure vegetative mycelium induced by sterilized water(exchanging water per three hours),and sporangium were produced from the seventh hour after treatment.The research determined DDRT-PCR optimization system for amplifying P.sojae RNA,and selected 30 pairs of primer combinations,finale,obtained 90 stable differential fragments.The 39 positive differential fragments were tested by reverse northern hybridization,and false positive rate was 56.7%.we obtained 36 differential fragments sequences after cloning and sequencing;the homologous alignment were tested by Blast software in Genbank database.The results showed that 9 fragments did not have matched homologous sequences,the remaining 27 fragments had high homology with many ESTs of Phytophthora sojae,Phytophthora infestans,Phytophthora brassicae,Phytophthora nicotianae in the processes of stress,growth of mycelium and spores,infection and propagation etc.; only 9 of 27 fragments had homology with known functional gene and protein sequences.Differential fragments of AR₇-296 and GR₇-292 had 74%homology with dolichylphosphate mannosyltransferas EDL06543.1,it might encode dolichylphosphate mannosyltransferase related in the synthesis of cell wall,and regulated for transformation of mannose to provide for the formation of cell wall.The differential fragment GR₁₉-312 had 80%homology with the gene NM125543.3 of arabidopsis autophagy 3,it might encode autophagy gene,and participate in the autophagy process of Phytophthora sojae to provide energy for the apical growth of mycelium and formation of sporangium.The differential fragment CR₁₅-282 had 75%homology with the gene NM₀01030328.2 of xenopus tropicalis Rab5,it might encode Rab5 gene,and regulated vesicles formation to provided vesicles for the apical growth of mycelium and formation of sporangium. The differential fragment CR₁₅-290 had 70%homology with assumed senescence related protein BAB33421.1,it might encode senescence related protein to regulate the senescence process of Phytophthora sojae by ubiquitin ligase E3.The differential fragment GR₁-373 had 36%homology with diacylglycerol acyltransferase XP₈21103.1,it might participate in the synthesis of oil.The differential fragment AR₁₅-258 had 30%homology with ABC carrier protein XP₆43503.1,it may be related to toxic metabolites outside cells.The differential fragment CR₄-289 had 83%homology with ribonucleoprotein binding protein XM001654574.1,it might encode ribonucleoprotein binding protein,but the function of this protein remained unclear,the differential fragment GR₁₉-366 had 72%homology with hypothesis protein XP₀01618281.1,it might encode hypothesis protein,but the function of this protein remained unclear.