Quantitative real-time PCR (qRT-PCR) for miRNAs only required a forward primer to be designed for the individual miRNA. The reverse primer was a universal primer provided with the miScript SYBR Green PCR Kit (Qiagen). All known chicken miRNA sequences were obtained from miRBase [34]. Oligonucleotide primers for these miRNAs were designed using full-length mature miRNA sequences. Primers were synthesized by Genotech (Daejeon, South Korea) (see Additional file 3).
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