微生物学微生物学 (43).docVIP

Hi,everyone,inthissection,wewilltalkaboutAmestest.Afterwatchingthisvideo,youshouldbeabletotelltheprincipleofAmestestanditsapplication.

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Mutagenicchemicalsmayincreasethepossibilityofhavingcancer.Findingoutthemutagenicityofachemicalsubstanceisasignificantpartoftestingthesafetyofasubstance.Nowadays,Amestesthasbecomeavalidproceduretoidentifycarcinogens.Inthe1970s,BruceAmes,developedthisexperimenttoassessthepotentialcarcinogeniceffectofchemicalsbyusingaparticularstrainofSalmonellatyphimurium.Therefore,thistestisalsocalledSalmonellatyphimuriumreversemutationassay.

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Firstly,let’stakealookattheprincipleoftheAmestest.OneofthecommonlyusedstrainsofbacteriaisSalmonellaTyphimurium.Itcarriesamutationinagenethatcontainshistidine,thismutanttermedashis-strain,whichrequireshistidineingrowthmedia.Whenachemicalmutagenwithpotentialcarcinogeniceffectisaddedintothegrowthmedia,reversemutationmaybeoccur;thehis-strainwouldbeexpectedbeingtransformedintohis+strain,whichrequiresnohistidineingrowthmedia.Therefore,thenumberofcoloniesonthegrowthmediawithminimalhistidineaddedwouldvarydependingontherateofreversemutation,inotherwords,themutagenicityofthetestedchemical.

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Next,let’stakealookattheproceduresofAmestest.Firstly,atestsuspensionofSalmonellatyphimuriumshouldbepreparedandplacedinaplainbuffercontainingthechemicaltobetestedfor.Secondly,addafewamountsofhistidinetoinitiatebacterialgrowth.Onlybacteriamutatedintohis+hastheabilitytosynthesizehistidine,suchbacteriawillformacolony.Thirdly,getacontrolsuspensionofSalmonellaTyphimuriumminusthetestchemicals.Fourthly,incubateforapproximately20minutesat37°C.Fifthly,scatterthesuspensionontheagarplate,andsixthly,incubatetheplatefortwodaysat