关键词: 西方蜜蜂（Apis mellifera L.）, sRNA富集, miRNA文库, 直接克隆法, 高通量测序

Abstract:

【Objective】 The aim of this study is to separate and enrich sRNAs from honey bee (Apis mellifera L.), and construct a library to test whether these sRNAs could meet requirements of high-throughput sequencing research. 【Method】 sRNAs (15-40 nt) from the queen, worker and drone bees at different development stages were separated and enriched, then performed reverse transcription on adapter ligated sRNAs followed by PCR amplification. After constructing the cDNA library, blue-white selection was performed and 288 single clones were selected for sequencing and analysis. 【Result】 214 high quality sequences of 15-39 bp, including 65 sme-miR-71c (mir-2), 5 ncRNAs containing tm-RNA, intron_ghI and 5.8S rRNA, 28 tRNAs, 33 siRNAs and other sRNAs, 1 CDS, and 82 not mapped sequences, were obtained. 【Conclusion】 This result indicates that the sRNA enrichment is effective and the samples we prepared could be used for high-throughput sequencing.

Key words: honeybee (Apis mellifera L.), sRNA enrichment, miRNA library, direct cloning method, high-throughput sequencing methods