RECOMBINANT RNA PHAGE Q-BETA CAPSID PARTICLES SYNTHESIZED AND SELF-ASSEMBLED IN ESCHERICHIA-COLI

The Escherichia coli RNA phage Q beta coat protein-encoding gene (C) w as amplified from native Q beta RNA using a reverse transcription-PCR technique. Gene C contains sequences coding for both the 133-amino aci d (aa) Q beta coat protein (CP) and the 329-aa read-through protein (A 1) consisting of CP and an additional 196-aa C-terminal sequence, sepa rated from CP within the C gene by an opal (UGA) stop condon. Primers ensuring the natural environment for gene C, especially within the rib some-binding site, and supplying C with unique restriction sites at bo th ends have been prepared. An amplified 1062-bp PCR fragment was posi tioned under the control of the strong E. cole trp promoter (P-trp) wi thin a pGEM-derived plasmid. The synthesis of gene C products was conf irmed electrophoretically and immunologically. An immunodiffusion test with anti-Q beta phage antibodies and electron microscopy evaluation of the purified recombinant products showed that when expressed, the Q beta C gene was responsible for high-level synthesis and correct self -assembly of Q beta CP monomers into capsids indistinguishable morphol ogically and immunologically from Q beta phage particles, which we pla n to use as surface display vectors.