Fig. 5. RNAi-mediated knockdown of Aaslif and AaiCAT2 inhibits amino acid stimulation of the vg gene. 1 day-old mosquitoes were injected with 0.6-1.0 µg of the following dsRNAs: the non-coding region of a control bacterial gene (MAL), the coding region of neutral amino acid transporter (NAT), the coding region of the Aaslif gene (Slif), the coding region of the AaiCAT2 gene (iCAT2), or a 1:1 mixture of both Aaslif and AaiCAT2 dsRNAs. Mosquitoes were allowed to recover for 5 days. Fat bodies from these mosquitoes were then dissected and cultured in either the presence or absence of amino acids (AAs) for 6 h. Total RNA was isolated from three groups of six fat bodies per treatment. cDNA was synthesized from equal amounts of DNase I-treated total RNA. (A) Gene expression was analyzed using vg-specific real-time PCR primers. Data were normalized by real-time PCR analysis of actin levels in the cDNA samples. Values are means ± s.e.m. of triplicate samples. (B) Knockdown of transporter genes was confirmed by RT-PCR analysis of the same cDNA used for the analysis in A.