沼泽红假单胞菌phbc-hupl缺失突变株构建及产氢特性研究-construction and hydrogen production characteristics of rhodopseudomonas palustris phbc - hupl deletion mutant.docxVIP

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沼泽红假单胞菌phbc-hupl缺失突变株构建及产氢特性研究-construction and hydrogen production characteristics of rhodopseudomonas palustris phbc - hupl deletion mutant.docx

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沼泽红假单胞菌phbc-hupl缺失突变株构建及产氢特性研究-construction and hydrogen production characteristics of rhodopseudomonas palustris phbc - hupl deletion mutant

TheEmrcassettewasclonedintothe‘pSUP202-phbC’,generatingthesuicidegenevectorpYMT.ThepYMTwastransformedintoE.coliS17-1,obtainingthedonorbacteria.Thedonorwasmixedwiththeoriginalstrain(receptorbacteria)R.palustrisCQU01.ThecorrectintegrationofthemutantswasconfirmedbyPCRtestandsubsequentsequenceanalysis.ThesinglemutantstrainR.palustrisCQU013withphbCknockedoutanddoublemutantstrainR.palustrisCQU014withphbC-hupLknockedoutwereobtained.Underthesameconditions,thebacterialgrowthcurvesandhydrogenproductioncharacteristicsofthemutantstrainsshowedthatthebiologicalcharactersofthemutantstrainshadsomedifferencewiththewildstrainR.palustrisCQU01.Twomutantstrainsofbacteriaincreasedmainlyinlessthan120h,andthebacteriawasnotsignificantlyincreased.Butthewildstrainofthebacteriaincreaseduntil168h,andthenbegantodecline;Mutantstrainhaslogarithmicgrowthphaselongerthanthewildstrain.Butthemaximumbiomasswassignificantlylowerthanthewildstrain.Themutantstrainsmayconsumealargenumberofenergy,sostrainbiomassislowerthanthewildstrain.Hydrogenproductionresultsalsoshowthatthetwomutantstrainshavetheremarkableenhancementofhydrogen.Thesamefermentationconditions,ThetotalH2yieldofR.palustrisCQU013andR.palustrisCQU014increasedby31%and76%respectivelycomparingtowildstrain,whichupto454mLand604mLperliterfermentationbrothin216hours.BoththephbCgeneandhupLgeneaffectedtheyieldofhydrogenproduce.Inparticular,thedoublemutantstrainisexpectedasaneffectiveengineeringstrainforphotosynthetichydrogenproductionandmicrobialbiologicaltreatmentofindustrialwastewater.ThestudieshaveshownthathydrogenproductioncapacityofdoublemutantstrainismorethanthephbCgeneandthesinglehupLgenemutant.Twogenesareinvolvedinthemetabolismofphotosynthetichydrogenproduction.Doublemutantcanimprovethehydrogenproductionsignificantly.Buttheresultofbacterialmetabolicpathwaysvariabilityanddiversity,theremaybeothermetabolicpathways.Sotheincreaseislimited.OnR.palustrishydrogenproductioncapabilities,wehavetounderstandthehydrogenmetabolicpathwayandthemechanism.Keywords:R