IÕve just started to perform site-directed mutagenesis. Could
I use the pBluescript(pBS) phagemid and CJ236 bacterial
strain with VCSM13 helper phage to replicate my single-
stranded(ss) pBS DNA? Second question is someone only gives
me a ssDNA pBS phagemid (inserted gene) with dUTP. Could I
replicate this DNA in CJ236 bacteria without uridine and then
replicate it in DH5-alpha bacteria to get my double-
stranded(ds) DNA without dUTP? If it can work, is there any
possible to have mutation sites in the pBS phagmid in this
procedure? Third question is someone gives the protocols of
manipulate the ssDNA with VCSM13 helper phage, but this
protocol does not add any kanamycin in culture process. Does
it work to let me to get ssDNA of pBS phagmid?
I appreciate any advice on this matter.