Small RNAs (sRNAs) are vital regulators of gene expression and involved in various biological processes. Among them, microRNAs (miRNAs) and phased small interfering RNAs (phasiRNAs) have been well defined and studied in the past decades. A bunch of scripts or pipelines were developed to annotate miRNAs and phasiRNAs. However, some computational annotations are rough and without careful manual check, resulting in low quality annotation. In this study, 19 public strawberry (Fragaria vesca) sRNA sequencing data from nine different tissues were collected to annotate miRNAs and PHAS loci in F. vesca. After bioinformatics analysis and careful manual checking, 167 known miRNAs, 27 miRNA*s with notable abundance, 54 novel miRNAs were accurately annotated. The terms of two miRNAs were corrected from miR477b and miR5225 using miRN47 and miR3627h, respectively. Besides 21 nucleotides (nt) miR390, eleven miRNAs with a length of 22-nt are in charge of triggering the biogenesis of 21-nt phasiRNAs from 110 PHAS loci in strawberry. In particular, we found several PHAS loci were targeted by two different miRNAs (similar to the “two-hit” model) and the phasiRNA generating region located between two target sites. We speculate that one target site is in control of triggering phasiRNA biogenesis and the other target site define the boundary of the region of phasiRNA biogenesis, which likely provide an accurate way for phasiRNA generation. Overall, we provided a comprehensive and accurate annotation of miRNAs and PHAS loci in the F. vesca genome.

小RNA（sRNA）是基因表达的重要调节剂，并参与各种生物过程。其中，在过去的几十年中，对microRNA（miRNA）和阶段性小干扰RNA（phasiRNA）进行了很好的定义和研究。开发了许多脚本或管道来注释miRNA和phasiRNA。但是，某些计算注释很粗糙且没有仔细的手动检查，导致注释的质量较低。在这项研究中，收集了来自9个不同组织的19种公共草莓（Fragaria vesca）sRNA测序数据，以注释F. vesca中的miRNA和PHAS基因座。。经过生物信息学分析和仔细的手动检查，准确地注释了167个已知的miRNA，27个显着丰富的miRNA *，54个新颖的miRNA。使用miRN47和miR3627h分别从miR477b和miR5225纠正了两个miRNA的术语。除了21个核苷酸（nt）miR390外，还有11个长度为22-nt的miRNA负责触发草莓中110个PHAS位点21-nt phasiRNA的生物发生。特别是，我们发现了几个PHAS基因座被两个不同的miRNA（类似于“两次命中”模型）和位于两个靶位点之间的phasiRNA产生区域所靶向。我们推测一个目标位点在触发phasiRNA生物发生的控制中，而另一个目标位点定义了phasiRNA生物发生区域的边界，这可能为phasiRNA产生提供了一种准确的方法。总体而言，我们提供了F. vesca基因组中miRNA和PHAS基因座的全面准确的注释。