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商品描述
Themyo-InositolAssayKitisareliableandaccurateenzymaticUV-methodforthespecificmeasurementandanalysisofmyo-inositolinfoods,dairyproductsandvariousothermaterials.
Phyticacidcontentofsampleswithverylowlevelsoffreemyo-inositolcanalsobedeterminedusingK-INOSL.Thiscanbeachievedbymeasuringtheamountofmyo-inositolreleasedafterde-phosphorylationofphyticacidwiththeenzymesphytaseandalkalinephosphatase,asusedwiththeMegazymePhyticAcid/TotalPhosphorusAssayKit(K-PHYT).
Notsuitableforthedeterminationofmyo-inositolinbabyformula.
L-LysineproductionindependentoftheoxidativepentosephosphatepathwaybyCoryNEBacteriumglutamicumwiththeStreptococcusmutansgapNgene.
Takeno,S.,Hori,K.,Ohtani,S.,Mimura,A.,Mitsuhashi,S.&Ikeda,M.(2016).Metabolicengineering,37,1-10.
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WehaverecentlydevelopedaCorynebacteriumglutamicumstrainthatgeneratesNADPHviatheglycolyticpathwaybyreplacingendogenousNAD-dependentglyceraldehyde3-phosphatedehydrogenase(GapA)withanonphosphorylatingNADP-dependentglyceraldehyde3-phosphatedehydrogenase(GapN)fromStreptococcusmutans.StrainRE2,asuppressormutantspontaneouslyisolatedforitsimprovedgrowthonglucosefromtheengineeredstrain,wasproventobeahigh-potentialhostfofL-lysineproduction(Takenoetal.,2010).Inthisstudy,thesuppressormutationwasidentifiedtobeapointmutationinrhoencodingthetranscriptionterminationfactorRho.StrainRE2stillshowedretardedgrowthdespitethemutationrho696.OurstrategyforreconcilingimprovedgrowthwithahighlevelofL-lysineproductionwastouseGapAtogetherwithGapNonlyintheearlygrowthphase,andsubsequentlyshiftthiscombination-typeglycolysistoonethatdependsonlyonGapNintherestofthegrowthphase.Toachievethis,weexpressedgapAunderthemyo-inositol-inducIBLepromoterofiolT1encodingamyo-inositoltransporterinstrainRE2.TheresultingstrainRE2AiolwasengineeredintoanL-lysineproducerbyintroductionofaplasmidcarryingthedesensitizedlysC,followedbyexaminationforcultureconditionswithmyo-inositolsupplementation.Wefoundthatasahigherconcentrationofmyo-inositolwasaddedtotheseedculture,thefollowingfermentationperiodbecameshorterwhilemaintainingahighlevelofL-lysineproduction.ThisfinallyreachedafermentationperiodcomparabletothatofthecontrolGapAstrain,andyieldeda1.5-foldhigherproductionratecomparedwithstrainRE2.ThetranscriptlevelofgapA,aswellastheGapAactivity,intheearlygrowthphaseincreasedinproportiontothemyo-inositolconcentrationandthenfelltolowlevelsinthesubsequentgrowthphase,indicatingthatimprovedgrowthwasaresultofincreasedGapAactivity,especiallyintheearlygrowthphase.Moreover,blockadeofthepentosephosphatepathwaythroughadefectinglucose6-phosphatedehydrogenasedidnotsignificantlyaffectL-lysineproductionintheengineeredGapNstrains,whileadrasticdecreaseinL-lysineproductionwasobservedforthecontrolGapAstrain.DeterminationoftheintracellularNADPH/NADP+ratiosrevealedthattheratiosintheengineeredstrainsweresignificantlyhigherthantheratioofthecontrolGapAstrainirrespectiveofthepentosephosphatepathway.TheseresultsdemonstratethatourstrainengineeringstrategyallowsefficientL-lysineproductionindependentoftheoxidativepentosephosphatepathway.
Regulationofmyo-inositolbiosynthesisbyp53-ISYNA1pathway.
Koguchi,T.,Tanikawa,C.,Mori,J.,Kojima,Y.&Matsuda,K.(2016).InternationalJournalofOncology,48(6),2415-2424.
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Inresponsetovariouscellularstresses,p53exertsitstumorsuppressiveeffectssuchasapoptosis,cellcyclearrest,andsenescencethroughtheinductionofitstargetgenes.Recently,p53wasshowntocontrolcellularhomeostasisbyregulatingenergymetabolism,glycolysis,antioxidanteffect,andautophagy.However,itsfunctionininositolsynthesiswasnotreported.Throughamicroarrayscreening,wefoundthatfivegenesrelatedwithmyo-inositolmetabolismwereinducedbyp53.DNAdamageenhancedintracellularmyo-inositolcontentinHCT116p53+/+cells,butnotinHCT116p53-/-cells.Wealsoindicatedthatinositol3-phosphatesynthase(ISYNA1)whichencodesanenzymeessentialformyo-inositolbiosynthesisasadirecttargetofp53.Activatedp53regulatedISYNA1expressionthroughp53responseelementintheseventhexon.EctopicISYNA1expressionincreasedmyo-inositollevelsinthecellsandsuppressedtumorcellgrowth.KnockdownofISYNA1causedresistancetoadriamycintreatment,demonstratingtheroleofISYNA1inp53-mediatedgrowthsuppression.FurThermore,ISYNA1expressionwassignificantlyassociatedwithp53mutationinbladder,breastcancer,headandnecksquamouscellcarcinoma,lungsquamouscellcarcinoma,andpancreaticadenocarcinoma.Ourfindingsrevealedanovelroleofp53inmyo-inositolbiosynthesiswhichcouldbeapotentialtherapeutictarget.
Colourimetricmethodforthedeterminationofmyo-Inositol
invarioussamplematrices
Principle:
(myo-inositoldehydrogenase)
(1)myo-Inositol+NAD+→
2,4,6/3,5-pentahydroxycyclohexanone+NADH+H+
(diaphorase)
(2)INT+NADH+H+→NAD++INT-formazan
Kitsize: * 50assays
* Thenumberofmanualtestsperkitcanbedoubledifallvolumesarehalved.
ThiscanbereADIlyaccommodatedusingtheMegaQuantTM Wave
Spectrophotometer(D-MQWAVE).Method: Spectrophotometricat492nm
Reactiontime: ~25min
Detectionlimit: 0.8mg/L
Applicationexamples:
Animalfeeds,food,babymilkformulationandothermaterials
Methodrecognition: Novelmethod
Advantages
- Verycosteffective
- Allreagentsstablefor>2yearsafterpreparation
- Onlyenzymatickitavailable
- Rapidreaction
- Mega-Calc™softwaretoolisavailablefromourwebsiteforhassle-freerawdataprocessing
- Standardincluded
