" I want to use the gateway system of invitrogen to = minimize the
workload of subcloning our cDNAs in various expression = vectors.
using the system I have to clone our cDNAs in an = appropriate pEntr
vector. I used pEntr1a for this but I do not get any = colonies on
kanamycin plates using homemade DH5alpha or Invitrogen = Top10 cells.
did remove the ccdB sequence before putting in our cDNA. = I also
cutting out the ccdB sequence, religating the empty = vector and
transforming bacteria with this. I get only a very limited = amount of
colonies from transformed Top10 cells and none on our own = competent
What protocol are you following to enter the plataform? = Are you
making the BP reaction to clone your DNA into the entry vector?