Bead homogenizer

To measure mRNA expression, total RNA in cultured lung cancer cells was prepared by using TRIzol (Invitrogen, Carlsbad, CA, USA) and isopropanol (Sigma-Aldrich, St. Louis, MO, USA). To measure mRNA expression in A549-driven tumours, gastrocnemius muscle and epididymal white adipose tissue (eWAT), sacrificed tissues were frozen with liquid nitrogen and frozen tissues were pulverized by using dry ice and liquid nitrogen. Frozen tissues (30~40 mg) were homogenized with 1 mL of TRIzol (Invitrogen, Carlsbad, CA, USA) by using a bead homogenizer (Benchmark Scientific, Sayreville, NJ, USA) to extract total RNA. The purity of RNA was assessed by using a spectrophotometer (BioTek, Winooski, VT, USA) at 260 and 280 nm and a ratio of ~1.8 is accepted for measurement of gene expression. The reverse transcriptase and cDNA synthesis kit (Applied Biosystems, Foster City, CA, USA) was used for cDNA synthesis. SYBR Green qPCR mixture (Applied Biosystems, Foster City, CA, USA) was used for qRT-PCR and experimental Ct values were normalized and calculated to 36B4 (ribosomal protein subunit P0, RPLP0) or β-actin gene. Detailed primer sequences for qRT-PCR are described in Table 1.

A total of 279 female house flies were used for the PASA tests, and 18 randomly selected flies employed for the reverse transcriptase-PCR (RT-PCR) and Sanger sequencing. For genomic DNA (gDNA) extraction, the thorax of each individual house fly was excised and then homogenized using a bead homogenizer (Benchmark Scientific, Sayreville, NJ, USA). DNA was extracted from the homogenate using the DNeasy Blood & Tissue Kit (Qiagen, Hilden, Germany) following the manufacturer's recommended protocol. For the RT-PCR, total RNA was extracted from the thorax using the RNeasy Mini Kit (Qiagen) following the manufacturer’s protocol. The concentration of DNA and RNA in each sample was assessed using a NanoDrop 2000 UV spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA). The extracted DNA and RNA samples were preserved in a freezer at − 20 °C.