Activation of Akt Signaling Reduces the Prevalence and Intensity of Malaria Parasite Infection and Lifespan in Anopheles stephensi Mosquitoes
Figure 1
Generation of the CP-myr-AsteAkt-HA transgenic mosquito line and protein and transcript expression profile of the transgene in adult females.
A. Schematic of the construct genetically engineered into A. stephensi mosquitoes. See text for a description of the construct. B. Comparison of transgenic (TG) and non-transgenic (NTG) siblings. Top panel: non-transgenic (left) and transgenic fourth instar larvae (right) under white light. Middle panel: non-transgenic and transgenic mosquitoes under fluorescence and a DsRed filter. Bottom panel: merge of top and middle panels. C. Two transgene-specific primer sets were used to amplify the transgene from the genomic DNA of transgenic and non-transgenic siblings. Primers to AsteActin were used to verify the integrity of the DNA. D. Total RNA was isolated from the midguts or carcasses (i.e., entire body minus midgut) of both transgenic (TG) and non-transgenic (NTG) mosquitoes and converted into cDNA. Transgene specific primers were used to amplify myr-AsteAkt. Primers to AsteActin were used to verify the integrity of the cDNA. E. Total protein was isolated from the midguts or carcasses of transgenic and non-transgenic mosquitoes, separated electrophoretically on a 12% SDS-PAGE gel. Proteins were blotted and then probed with anti-HA antibody or anti-GAPDH antibody to assess protein loading.