Dear all,
Has anyone any (even the slightest) experience in expression of a cloned
protamine gene (mine derived from salmon) into an E. coli or Baculovirus
system?
At the moment we are unsuccesful in its expression and think it is a 'toxic'
peptide in the E. coli strains used. (pET 21a vector, BL21(DE3) and
BL21(DE3)pLysS E. coli strains). Also when using the Bac-to-Bac baculovirus
kit from Gibco-BRL, the recombinant bacmid does not appear to remain stable at
the E. coli stage and wild type revertants are very hard to avoid, which
subsequently, after transfection into Sf9 cells, are overproduced in the
viral stock.
Any tips please ???
Thanks,
GcR
___________________________________
Gert Roosen
PhD. Student
Royal Liverpool University Hospital
Duncan Building
Prescot Street L69 3BX Liverpool, UK.
+ 44 (0)151 706 4326 (tel)
+ 44 (0)151 706 4311 (fax)
gcr at liv.ac.uk (email)
