Bromothymol-Blue-sodium-生命科学试剂-MCE

Hotline:400-820-3792Inhibitors • ScreeningLibraries • Proteinswww.MedChemEBromothymolBluesodiumCat.No.:HY-D0012ACASNo.:34722-90-2分子式:C₂₇H₂₇Br₂NaO₅S分子量:646.36作用靶点:Others作用通路:Others储存方式: 4°C,sealedstorage,awayfrommoistureandlightInsolvent:-80°C,6months;-20°C,1month(sealedstorage,awayfrommoistureandlight)BIOLOGICALACTIVITY生物活性BromothymolBluesodiumsalt是一种pH指示剂，是测定弱酸弱碱时使用的一种指示剂，其酸性时为黄色，碱性时为蓝色。储存方式：避光。体外研究AfirstcharacterizationandcomparisonisdonebydevelopinganeasyanddirectmeasurementmethodbasedonapHindicatorsystemusingBromothymolBlue(BTB)astheindicatorandpotassiumphosphateasthebuffer.ApH-shiftassayisdevelopedonthebasisofBromothymolBlueasthepHindicatorandpotassiumphosphateasthebuffercomponent[1].ThreepHindicatorsaretestedforthedirectdeterminationof2-2-keto-L-gulonicacid(2-KLG)productiononaplate.TheresultsshowthatBromothymolBlueissuperiortotheothertwoindicatorsintermsoftheobviouscolorchangeandasuitablepHrange(bluetoyellowatpH6.5-7.5).UpontheadditionofaBromothymolBluesolution(0.1%,w/v)toanagarplate,zonessurroundingcoloniesofK.vulgare07mutantschangetheircolorfrombluetoyellowbecauseK.vulgare07mutantsrelease2-KLGonagarplates,therebyacidifyingsurroundingareasaroundcolonies[2].PROTOCOLKinaseAssay[1]Forallthreeenzymes,anexpressioninthe96-deepwellscaleisperformed.Therefore,electrocompetentE.coliArcticExpress(DE3)cellsaretransformedwiththecorrespondingplasmid.SingleclonesarepickedusingtheCP7200ColonyPickerandtransferredto96-deepwellplatesfilledwith1.2mLautoinductionmediabyaMicroFloSelectdispenser.Afterincubation(36h,37°Cat1,000rpm),furtherprocessingisdonemanually.First,100μLofcellcultureistransferredintoa96-wellplate(U-shapedbottom)andharvestedbycentrifugation(4,570rpm,10minatRT)whilethesupernatantisdiscarded.Thefrozenpellets(1hat-20°C)1/2 MasterofBioactiveMolecules—您身边的抑制剂大师www.MedChemEarethawedatroomtemperatureforonehourtoimprovecelllysis.Lysisiscontinuedbytheadditionof30μLlysisbuffer(3h,1,000rpm,37°C)containing2mMKPi,pH7.0,2mMMgCl2,10μg/mLDNaseI,100μg/mLlysozyme.Next,120μLbuffer(2mMKPi,pH7.0)isaddedfollowedbycentrifugation(3,000rpm,15minatRT).Forthephotometricmeasurement,20μLofthecrudeextractistransferredtoa96-wellplate(F-shapedbottom)andthereactionisstartedbyadding180μLmastermixtogiveafinalvolumeof200μL(2.5mMKPi,pH7.0,2mMMgCl2,25μg/mLBromothymolBlue(BTB)and5mMketo-deoxy-D-glucarate).Themeasurementsarecarriedoutfor60minat2-minintervals.Dependingontheenzyme,differenttimewindowsareusedfortheactivitycalculation[1].MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.REFERENCESPickA,etal.Identificationandcharacterizationoftwonew5-keto-4-deoxy-D-GlucarateDehydratases/Decarboxylases.BMCBiotechnol.2016Nov17;16(1):80.YangW,etal.AplatemethodforrapidscreeningofKetogulonicigeniumvulgaremutantsforenhanced2-keto-l-gulonicacidproduction.BrazJMicrobiol.2017Jul-Sep;48(3):397-402.McePdfHeightCaution:Producthasnotbeenful