Analyses of the proteins expressed by the BVboostFG system

Copyright information:Taken from "A multipurpose vector system for the screening of libraries in bacteria, insect and mammalian cells and expression "Nucleic Acids Research 2005;33(4):e42-e42.Published online 24 Feb 2005PMCID:PMC549581.© The Author 2005. Published by Oxford University Press. All rights reserved () Western blot of the expressed avidin detected with an avidin antibody. M, molecular weight markers; control avidin marked with an arrow. T, sample of total proteins; P, periplasmic fraction; I, insoluble fraction. The upper faint band represents ompA secretion signal-containing avidin whereas the lower band represents the fully processed avidin. Control avidin has a higher molecular weight because it contains a carbohydrate side chain (). () Deglycosylation analysis of purified VEGF-A produced in insect cells. N indicates a sample not treated with endo H and T indicates a sample deglycosylated with the enzyme. () A dot-blot analysis of SEAP secreted into insect cell medium. The uppermost dot is 200 μl of medium diluted 1:10, whereas the middle dot is a positive control containing 300 ng of SEAP. The lowest dot is a negative control. () and () show purified sKDR(1–7) (silver staining) and VEGF-D (Coomassie staining) proteins in SDS–PAGE gel, respectively. The proteins were purified to milligram amounts with no apparent contaminants which indicate high purity. The lower bands seen in VEGF-D preparation are different glycosylation forms of the protein. () and () show immunoblots of HIV-integrase produced in and insect cells, respectively. Most of the protein produced in bacteria was degraded whereas protein produced in insect cells was intact. Proteins were detected with an integrase-specific antibody.