Transcriptome analysis of Gossypium hirsutum flower buds infested by cotton boll weevil (Anthonomus grandis) larvae-Sample8 cotton control
Sequencing libraries were prepared using NEBNext Ultra Directional RNA Library Prep Kit (New England Biolabs, MA), which included Poly-A-containing mRNA isolation from 5 µg total RNA, using two purification rounds of poly-T oligo-attached magnetic beads, and fragmentation by sonication. First strand cDNA was generated using reverse transcriptase and random primers. Following the second strand cDNA synthesis and adaptor ligation, 400 bp cDNA fragments were isolated using gel electrophoresis and amplified by PCR. The products were loaded onto an Illumina HiSeqTM 2000 instrument and subjected to 200 cycles of paired-end (2 × 100 bp) sequencing.