I am creating scFv phage display libraries using the VCSM13 helper phage (Agilent). It isn’t clear to me whether protein GIII can be cleaved by trypsin in this phage. To my knowledge its phenotype isn’t published, or at least I can’t find any clear information on this issue.
I have incubated 10^10 phage particles in PBS +/- trypsin, infected TG1 cells and titrated them out on agar plates with kanamycin (VCSM13 confers kanamycin resistance). I saw equal numbers of colonies, regardless of trypsin digestion. Either this helper phage isn’t susceptible to trypsin, or I picked a defective plaque prior to culturing the helper phage.
Has someone got experience with this phage?
Maybe this answer is a bit too late, but VCSM13 (or the pIII coat protein that confers its infectivity) is not supposed to be sensitive to trypsin. In fact, trypsin digestion is one of the methods to elute and retrieve antibody-displaying phage particles from the antigen-coated surface (Barbas et al. Phage Display: A Laboratory Manual, Cold Harbor Springs Laboratory Press). Incidentally, the pIII protein sequence of VCSM13 is identical to the wild-type M13 as well as other conventional helper phages such as M13K07, so they are trypsin-resistant too.
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Cell Biology
- Culture Cells
- Cell Line