title: GSM2564799: NSR_Testis; Anolis carolinensis; RNA-Seq
library_construction_protocol: Total RNA and small RNA was extracted from homogenized testis and/or brain tissue using TRIzol . For NSR-Seq libraries: First strand cDNA synthesis was performed using SuperScript III and the “not-so-random” forward primer set (5’ TCCGATCTCTNNNNNNN 3’), followed by second strand synthesis using exo- Klenow fragment and the “not-so-random” reverse primer set (5’ TCCGATCTGANNNNNNN 3’). In order to deplete ribosomal RNAs from the libraries, “not-so-random” primer sets were designed by pooling 749 unique hexamers with no perfect match to human cytoplasmic 18S and 28S rRNA and mitochondrial 12S and 16S rRNA transcripts.  Libraries were subsequently amplified by PCR using primers (5’ AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCTCT 3’ and 5’ CAAGCAGAAGACGGCATACGAGATCGGTCTCGGCATTCCTGCTGAACCGCTCTTCCGATCTGA 3’) and paired-end sequenced for 50x2 cycles on Illumina Genome Analyzer II according to manufacturers’ instructions. For small RNA libraries: RNA was ligated to 3′ and 5′ adaptors, gel purified after each ligation, reverse transcribed, and PCR amplified using Solexa sequencing primers. PCR product was gel purified, quantified, and sequenced for 36 cycles or 76 cycles on Illumina Genome Analyzer II according to manufacturers’ instructions. "Not-So-Random" RNA-Seq, small RNA-Seq
identifiers.namespace: GEO
identifiers.id: GSM2564799