Tnt coupled reticulocyte lysate system kit

In all the NAPPA assays, to check the deposition of the cDNA on the microarray surface, microarrays were blocked, and cDNA staining was carried out by incubating each microarray with 200 μl of picogreen solution diluted by 1:600 (v/v). Additionally, to observe in situ protein expression, arrays were blocked and incubated with IVTT system kit (TnT^® Coupled Reticulocyte Lysate System kit, Promega) and analyzed as previously described (23 (link), 24 (link)).

Total RNA was purified from whole larvae and adults by tissue homogenization and Qiazol reagent extraction. CG2145 and CG3303 ORFs were amplified, through the oligonucleotides A (5′-ATGCGCTGCCTGGCATTGAGC-3′), B (5′-TCAAATCTCCGGATAGGCGCTGCC-3′) and C (5′-ATGGACGCAGTGCAGCAGCAAAC-3′), D (5′-CTAGGAGCTGTCCGGATAGAC-3′), respectively, and cloned into the pGEX-4T1 vector, between the EcoRI and NotI sites. CG2145 and CG3303 [³⁵S]methionine-labeled proteins were produced by in vitro transcription and translation reactions using the TNT-coupled Reticulocyte Lysate System kit (Promega), according to manifacturer’s instructions.