BP25_3_egg
Poly-A RNA was enriched from 0.6-1.0 µg total RNA using NEXTflex Poly(A) Beads from Bioo Scientific. Strand-specific cDNA libraries were prepared using the Bioo Scientific NEXTflex Directional RNA-Seq Kit (dUTP Based) for Illumina following manufacturer instructions with a 12 min fragmentation time and 15 PCR cycles. Libraries were barcoded with Bioo Scientific adaptors so they could be pooled for sequencing. Library concentrations were quantified using a Qubit dsDNA High Sensitivity Assay Kit (Life Technologies), and library size was assessed using a Bioanalyzer High Sensitivity DNA chip (Agilent). Libraries were pooled into four groups at equal concentration and diluted to final pooled concentrations of 10 nM. Library pools were quantified using the Illumina-compatible kit and KAPA standards for real-time PCR by the W. M. Keck Center for Comparative and Functional Genomics at the Roy J. Carver Biotechnology Center (University of Illinois).