Functional identification of lncRNAs in sweet cherry (Prunus avium) pollen tubes via transcriptome analysis using single-molecule long-read sequencing

利用单分子长读长测序技术进行转录组分析,鉴定甜樱桃(Prunus avium)花粉管中lncRNA的功能

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Abstract

Sweet cherry (Prunus avium) is a popular fruit with high nutritional value and excellent flavor. Although pollen plays an important role in the double fertilization and subsequent fruit production of this species, little is known about its pollen tube transcriptome. In this study, we identified 16,409 transcripts using single-molecule sequencing. After filtering 292 transposable elements, we conducted further analyses including mRNA classification, gene function prediction, alternative splicing (AS) analysis, and long noncoding RNA (lncRNA) identification to gain insight into the pollen transcriptome. The filtered transcripts could be matched with 3,438 coding region sequences from the sweet cherry genome. GO and KEGG analyses revealed complex biological processes during pollen tube elongation. A total of 2043 AS events were predicted, 7 of which were identified in different organs, such as the leaf, pistil and pollen tube. Using BLASTnt and the Coding-Potential Assessment Tool (CPAT), we distinguished a total of 284 lncRNAs, among which 154 qualified as natural antisense transcripts (NATs). As the NATs could be the reverse complements of coding mRNA sequences, they might bind to coding sequences. Antisense transfection assays showed that the NATs could regulate the expression levels of their complementary sequences and even affect the growth conditions of pollen tubes. In summary, this research characterizes the transcripts of P. avium pollen and lays the foundation for elucidating the physiological and biochemical mechanisms underlying sexual reproduction in the male gametes of this species.

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