JeffLawrence(M02)1.Place2mloftheappropriatesterilemediumina13mmyellow-cappedculturetube.Ifmorecultureisneeded,placeupto5mlina16mmgreen-cappedculturetube.Largeamountofcultureshouldbemadeingrowthflasks,usingupto25%flaskvolumeinmedium(e.g.,nomorethan30mlina125mlflask,or250mlina1000mlflask).2.Touchasinglecolonywiththeendofanapplicatorstickandholditinyourprimaryhand(righthandifright-handed).3.Holdingthetesttubeinyouotherhandwithpalmandlowerfingers,removethecapofthetesttubewithyourthumbandindexfinger.Diptheinoculatedendoftheapplicatorintothesterilemediumand,ifnecessary,wipetheendontothesideofthetube.4.Brieflyflametheendofthetesttubeandreplacethecap.5.Alwaysinoculateadummytubewithnocellstoverifythatthemedium,tubes,andsticksweresterile.6.Ifinoculatinglargecultures,diluteanexistingculture1:100.7.Growculturefor12to18hours.After24-30hours,cellshaveenteredstationaryphaseandshouldnotbeusedforcriticalprocedures.8."Quicky"overnightscanbemadebyrescuinga2or3dayculture.Diluteculture1:10insterilemediumandgrowfor3hours.
