Differential Host Cell Gene Expression and Regulation of Cell Cycle Progression by Nonstructural Protein 11 of Porcine Reproductive and Respiratory Syndrome Virus
Figure 1
Establishment of MARC-nsp11 cells stably expressing PRRSV nsp11. (a) Incorporation of nsp11 gene in cellular DNA and identification of cell clones. A total of 7 clones, designated nsp11-a through nsp11-g, were obtained and screened for nsp11 sequence by PCR. Cellular DNA was extracted and PCR was performed using the primers described in Materials and Methods. The nsp11-a clone was chosen to conduct RT-PCR (b) and immune-blot (c) and designated as MARC-nsp11 cells. (b) Total cellular RNA was extracted from MARC-145 and MARC-nsp11 cells and subjected to DNase I treatment followed by RT-PCR or PCR. (c) Cell lysates were prepared from MARC-145 (lane 1), nsp11-gene transfected MARC-145 (lane 2), and MARC-nsp11 (lane 3) cells and were incubated with Protein A Sepharose beads and anti-rabbit nsp11-specific polyclonal Ab, followed by immunoblot using anti-FLAG monoclonal Ab.
