title: GSM8990923: Ctrl 3-r/-mRNA, Replicate 1; Aedes albopictus; RNA-Seq
library_construction_protocol: Total RNA was extracted from wAlbB-infected C6/36 cells using the miRNeasy Mini Kit (Qiagen, Hilden, Germany), with protocol adaptations. Samples were incubated at 37 °C in a heating block until completely thawed, ensuring that all salts were fully dissolved. After an additional 5 min incubation at room temperature, samples were mixed with 100 µl 1-Bromo-3-chloro-2-propanol (BCP, Tokyo Chemical Industry, Tokyo, Japan). The sample-BCP mixture was vigorously shaken for 15 seconds, followed by a 2–3 min incubation at room temperature. Phase separation was achieved through a 15 min centrifugation at 12,000 g, 4 °C. The upper aqueous phase, containing the nucleic acids, was retained, with 350 µl applied to the Qiagen column. The extract was also subject to DNase I treatment to eliminate contaminating DNA. Two samples were subjected to rRNA depletion using riboPOOLs (siTOOLs Biotech, Planegg, Germany) according to the manufacturer's instructions, using the maximum input of 5 µg RNA. Three riboPOOLs were employed: Aedes albopictus (dp-K012-47) and custom-designed riboPOOLs for Wolbachia and Aedes albopictus. The sequences of the riboPOOLs are proprietary knowledge of siTOOLs Biotech.