Maintenance of Probes in bacteria including Escherichia coli
来自 : 蚂蚁淘
- Plasmid(pUCseries)containinggenomicDNAfragmentsaremaintainedinE.colistrainDH5aTM.TheE.coliculturesareroutinelyculturedat37ConLuria-Bertani(LB)agaronorinLBbrothcontainingAmpicillin(30µg/ml)orCarbenicillin(50µg/mlbroth,100µg/mlagar).E.colistrainsareusuallypreservedinstabagarorglycerolformid-termstorageandlyophilizedforlong-termstorage.
StorageinStabAgar:MoststrainsofE.colicanbestoredforonetotwoyearsinstabagar.
- Useairtight,autoclavedvialscontaining2/3full(appr.1ml)ofmedium.
- Inoculateeachvialwithasinglecolony.
- Leavethecapofthevialsightlylooseandincubateovernightat37C.
- Sealthevialtightlyandstoredatroomtemperatureinadarkplace.
- Toreviveastoredstrain,streakontoLBplatecontainingappropriateantibioticsforasinglecolony.
StorageinglycerolSolution:MoststrainsofbacteriaincludingE.coli,andfungi(sporesispreferred)canbestoredforonetotwoyearsinglycerolsolutionat-20C.At-70Ctheycanbestoredalmostforlifetime.
- Prepare0.5mlofglycerolsolutionin2.0mlscrew-capvials,autoclave(15min,121C),cool,andstorethematroomtemperature.
- Transferonedrop(0.5ml)ofanovernight-growncultureintoavial.
- Voltex,storeat-20or-70C(-70Cstorageispreferredforlong-termmaintenance).
- Toreviveastoredstrain,streakontoLBplatecontainingappropriateantibioticsforasinglecolony.
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