关键词: 亚洲棉, 短绒突变体, 长链非编码RNA, 调控网络, 荧光定量PCR

Abstract: 【Objective】Long non-coding RNAs(lncRNAs) are a group of RNA molecules longer than 200 bp with no protein coding capacity, which are involved in various biological regulatory processes. In this study, we aim to analyze the RNA-sequencing data of two Gossypium arboreum isogenic lines, a fuzzless mutant and its wildtype, to identifying the lncRNA involved in early fuzz fiber development, providing a foundation for investigation the mechanism of fiber development. 【Method】We collected 0 DPA, 3 DPA and 5 DPA ovule and 8 DPA ovule and fiber from the G. arboreum fuzzless mutant GA0149 and its isogenic line GA0146 with normal fuzz and lint fibers, were used for RNA-seq to identify lncRNA and predict their target genes. Differentially expressed mRNA（DE-mRNA） and lncRNA(DE-lncRNAs) between the samples were identified. The KOBAS software was used to prediction the KEGG enrichment pathways which DE-lncRNAs targets were involved in. To ensure the quality of high-through sequencing, 25 DE- lncRNAs were selected for RT-qPCR detection. 【Result】We identified 15339 lncRNA-encoding transcripts that 11595 lncRNAs were located to intergenic regions, 2428 lncRNAs were classified as antisense lncRNAs, 350 were categorized as intronic lncRNAs and 966 were belonged to sense lncRNAs. Compared to mRNAs, lncRNAs in Asian cotton showed shorter exons and lower GC content. Most of lncRNAs had cis-regulatory effects on their neighboring mRNAs. We identified 1932 differentially expressed (DE) lncRNAs, with 8134 predicted DE-lncRNA target genes. Further analysis showed that 788 genes (mRNA) were differentially expressed (DE-genes) during four fiber development stages. KEGG enrichment pathways analysis showed that DE-target-mRNAs were mainly enriched in Plant hormone signal transduction and Protein processing in endoplasmic reticulum. Co-expression network analysis revealed that lncRNA (MSTRG.454250.3) and its associated target genes showed identical expression trends during four fuzz fiber development stages, while lncRNAs (MSTRG.454261.4) and its associated target genes showed contrary expression tendency, exhibiting dramatic higher expression in fuzzless GA0149 compared to wildtype GA0146. The results of RT-qPCR analysis confirmed the authenticity of our RNA-seq data.【Conclusion】A total of 26 specifically expressed lncRNAs were identified which related to cotton fuzz fiber development process. We further confirmed that these lncRNAs affected the fuzz fiber development by regulating the expression of indole-3-acetic acid-amido synthetase (Ga03G2421) and Auxin-responsive protein (Ga05G1344) in the plant hormone signal transduction pathway.

Key words: Gossypium arboreum, fuzzless mutant, long non-coding RNAs, regulation network, RT-qPCR