In vivoexpression of UNC93A.Cellular expression and localisation of GFP-UNC93A. The ORF of UNC93A was subcloned inframe into a fluorescent GFP tagged mammalian expression vector (EGFP-N3) and was transiently transfected into 293-T cells. 24 hours after the transfection, the cells were transferred to cover slips for a further 24-hour culture, followed by fixation and observed under fluorescent microscope. The fusion protein was localised on the membrane of the transfected cells and was abundant from 48 to 72 hours after the transfection. A ) cells observed under phase contrast, B ) cells observed under UV light, C ) merged image. D) Immunoprecipitation and immunoblotting of the EGFP-UNC93A fusion protein. Cell lysate from untransfected 293-T cells, 293-T cells transfected with EGFP vector, and 293-T cells transfected with EGFP-UNC93A (the EGFP + UNC93A lane) were immunoprecipitaed with GFP monoclonal antibody, resolved on an 10% SDS-PAGE gel, and Western blotted. A specific fusion protein of about 90 kDa was detected with GFP monoclonal antibody and anti-Mouse HRP in the cells transfected with EGFP-UNC93A, but not in the untransfected cells nor cells transfected with EGFP vector (indicated by arrow).