Name: P18856_2003_S11_p
Instrument: Illumina NovaSeq 6000
Strategy: ssRNA-seq
Source: TRANSCRIPTOMIC SINGLE CELL
Selection: PolyA
Layout: PAIRED
Construction protocol: L. sclopetarius adult female spiders were collected in the wild in a small habitat in Uppsala, Sweden. Taxonomic identity of the spider was verified by the Museum of Natural History, Stockholm, Sweden. The spiders were kept in big containers that allowed them to spin webs. They were fed with meal worms or Drosophila flies weekly and watered daily. In total 20 spiders were used for single cell sequencing. Method B: Due to problems with contaminating droplets of dope that made the separation of single cells challenging, a slightly modified protocol was used for the following ten spiders. These were processed as method A in Protocol 2, but with the exception that the duct was not removed, and the sac was cut and kept in PBS, pH 7.4 for 10 min to allow the dope to flow out. The pieces of glands were picked up and incubated in pre warmed trypsin-EDTA (Gibco, 0.5%) at 37°C for 2 min in a low-binding micro-centrifuge tube. The suspension was triturated with fire-polished glass Pasteur pipette for 1 min, incubated in trypsin-EDTA at 37°C for 2 min, and triturated again with fire polished glass Pasteur pipette for 3 min. The suspension was centrifuged at 300 23g for 3 min at 4°C and the pellet was resuspended in 200 μL DMEM containing 3% BSA. The suspension was strained through 40 μM pre-washed cell strainers into low-binding micro-centrifuge tubes. The strainer was further washed with 100 μL DMEM containing 3% BSA to reduce the loss of cells. The library preparation was done using 10X 3' GE kit on the 10X Chromium Single Cell 3' Platform following manufacturer's protocol (Dual Index 10X_3'_V3.1).