Preparation of North American Type II PRRSV Infectious Clone Expressing Green Fluorescent Protein
Figure 2
Synthesis of PRRSV-GFP mRNA in vitro and generation of the recombinant PRRSV containing gfp gene. (a) Electrophoresis of PRRSV-GFP mRNA (Lane 1) and PRRSV-GFP-Poly(A) mRNA (Lane 2) on 0.5% denaturing agarose gel, synthesized in vitro by T7 RNA polymerase using mMESSAGE mMACHINE mRNA Transcription kit. (b) GFP expression in Marc-145 cells infected for 72 h with the supernatant of the 3rd time of blind passage in BHK-21 cells originally transfected with PRRSV-GFP mRNA. (c) Identification of PRRSV expressing GFP by amplifying specific fragment by RT-PCR from total RNA from Marc-145 cells infected with PRRSV (Lane 1) and GFP-PRRSV (Lane 2). About 1100 bp fragment was amplified by RT-PCR with a pair of primers (F1 and R2).
