RNA-Seq of TSWV exposed F. fusca pupae
Total RNA was extracted from virus-exposed or non-virus exposed larvae (1st and 2nd instars), pupae (including pre-pupae), and female adults (up to 2 d old) (six treatments in total). For each treatment, about 35 individual thrips were pooled and treated as one experimental replicate. A total of three experimental replicates were used per treatment (N= 105 individual thrips per treatment). Total RNA was extracted using RNeasy plant mini kit using manufacturerês protocol (Qiagen). Subsequently, cDNA libraries were constructed at Georgia Genomic Facility of the University of Georgia. Illumina sequencing libraries were constructed using TruSeq RNA sample preparation kits using at least 1 µg of the total RNA. Briefly, mRNA was selected, fragmented, and first strand cDNA was synthesized using random primers and reverse transcriptase. Subsequently, Polymerase I and RNase H were used to make the second strand cDNA. An Illumina TruSeqLT adapter was ligated to the DNA fragments and PCR¾amplification was performed for a minimal number of cycles with standard Illumina primers to produce the final cDNA libraries for sequencing. Sequencing was performed at the University of Texas Health Science Center, at San Antonio, Texas on llumina HiSeq 2000 platform using v3 paired-end 100 cycle sequencing settings. Six RNA-seq libraries were included in one lane and a total of three lanes were used to sequence 18 libraries.