FOA培养基配置方法.doc

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1、5-FOA(5-FluorooroticAcid)MediumandAgarPlateRecipeFollowingrecipemakes1liter5-FOAplates:Step-1:Autoclave20gramsofagarin750mldoubledistilledwater.Step-2:Addfollowingsintoaglassbeaker,andbringto250mlwithdoubledistilledwater.AmmoniumSulfate                      5.0g  Yeastnitrogenbasewithoutam

2、inoacid  1.7gDextrose                                20g  Necessarynutritionalsupplements(seenote3)  Xg5-FOA                                    1.0g  Warmup5-FOAmixtureinStep-2at55-65°Conaheatplate,stiruntilitdissolves~30min-1h.Thesolubilityof5-FOAisverylowinwater;therefore,itisnecessaryto

3、warmupthesolution.Somemanufacturers,i.e.,USBiological,claimthat5-FOAisresistanttoautoclaving.Ididnottrytoautoclaveitmyself,butyoucantryit.After5-FOAcontainingmixtureinStep-2iscompletelydissolved,sterilizeitbyfilter.Placebothmeltedagarand5-FOAcontainingsolutioninawaterbath(50-60°C)foronehou

4、r.Finally,add5-FOAsolutionontomeltedagarandpouryourplates.PlatesmaybedriedatRTfor48hoursandmaybekeptat4°Cforlongtermstorage.Notes:1.DonotforgettoaddUracilintotheplatesat20mg/Loffinalconcentration(DonotaddtoomuchUracil!).2.NeveradjustthepHof5-FOAcontainingmedium.Onlyprotonatedformof5-FOAisp

5、ermeable.pHofyourplateswillbeveryacidicafteradding5-FOA.3.Donotaddmorenutritionalsupplementsthanitisnecessary;thisreducesefficacyof5-FOAplates.Forexample,youonlyneedtoaddHis,Leu,LysandUrasupplementsforstrainBY4742.Thesesupplementsmaybeaddeddirectlyfrom100xstocksolution.4.5-FOAisveryexpensi

6、vereagent.IfoundthatUSBiologicalhasthecheapestpricefor5-FOA.5.5-FOAishighlymutagenictoCandidaalbicans.Thismayalsobethecaseforbuddingyeast.Isuggestyoudonotleaveyeastcolonieson5-FOAplatesforlongerthanitisnecessary.WellingtonandRustchenko,Yeast,2005,Volume22,Issue1,Pages57-70).Othersourcesof5

7、-FOAisZymoResearchandSigma.TheUSBiologicalsellsthecheapest5-FOAreagent.Idonotknowifpurityof5-FOAisdifferentfromeachcompany.5-FOAfromUSBiologicalworkedgreatforme.Donotwasteyourmoney!5-FOA培养基与琼脂板配置方法下面是1L5-FOA平板的配方1：用750ml双蒸水灭20g琼脂2：将下列物质加入玻璃烧杯，同时加入250ml双蒸水硫酸铵5g