Whole transcriptome sequencing of the rhabdovirus-negative Sf9 cell clone 13F12
Cellular RNA was processed to generate libraries for mRNA sequencing following the Illumina TruSeq Stranded mRNA Sample Preparation Guide. In this methog, polyA-containing mRNA was purified from 1 g total RNA using poly-T oligo attached magnetic beads, fragmented and reverse transcribed into cDNAs. Double strand cDNAs were adenylated at the 3 ends and ligated to indexed sequencing adaptors, followed with brief amplification for 15 cycles. The quality of all cDNA libraries was assessed by Agilent 2100 Bioanalyzer using the DNA 1000 Kit (Agilent). One femtomole of the sequencing  libraries (median size ~260 nt) were denatured and loaded onto a flow cell for cluster generation using the Illumina cBot. The sample was loaded onto one lane of a HiSeq Rapid Run flow cell. Paired-end sequencing was carried out on HiSeq sequencer (Illumina) for 100 x 2 cycles. About 100 million 100-bp reads passed the preset filtering  parameters. The number of reads obtained after trimming in the CLC Genomics Workbench using a Phred score of 20 were 330,304,690.
Sf9-13F12