Figure 4.

Analysis of antibacterial activity of tDefA peptide isolated from transgenic mosquitoes. (A) Immunoblot analysis of peptide fractions used to assay the antibacterial activity. Protein samples obtained from transgenic 24 hPBM females of D1 line (TR), bacterially induced nontransgenic females of the kh^w host strain (BI), and nontransgenic 24 hPBM females of the kh^w host strain (NT) were isolated by using acid extraction and chromatography purification on Sep-Pak C18 cartridges and analyzed by immunoblotting. The analyzed samples were prepared from the 0% acetonitrile fraction in acidified water (0%) as a loading control, and the 40% acetonitrile fraction (40%), enriched for DefA peptide. (B) Aliquots from 40% fractions containing the major portion of defensin, isolated from transgenic (TR) and bacteria-induced (BI) females, were analyzed by the liquid growth inhibition assay using the Gram-positive bacteria, Micrococcus luteus. An equal aliquot of the 40% acetonitrile fraction isolated from nontransgenic 24 hPBM females of the host strain (NT) was used as a negative control. Data represent the mean of three independent experiments.