## 1 Introduction  
MicroRNAs (miRNAs) are small endogenous single-stranded non-coding small RNAs of 20-24 nucleotides (nts) that negatively regulate gene expression at the post-transcriptional level [1]. The biogenesis of miRNAs has been well elaborated in plants. Firstly, the long single-stranded pri-miRNAs (primary miRNAs) are generated by RNA polymerase II from the intergenic regions [2; 3; 4]. Secondly, the pri-RNAs are processed to generate 100-200 nt pre-miRNAs (precursor miRNAs) with stem-loop structures (hairpins) catalyzed by Dicer-like I enzyme (DCL1) in the nucleus, which creates a miRNA/miRNA* complex [5; 6]. Then these complexes are translocated into the cytoplasm and integrated with RNA-induced silencing complex (RISC), where the miRNA strand is incorporated with RISC and the miRNA* strand is usually degraded [7]. A large number of miRNAs have been discovered and identified from diverse plant species in the past decades. Traditional approaches including direct cloning and Sanger sequencing were widely employed for conserved miRNA identification in model plant species [8; 9]. However, some miRNAs are expressed in certain tissues or developmental stages and accumulate at relatively lower levels. The advent of next-generation high throughput sequencing technology provides a rapid and efficient opportunity for the identification of conserved miRNAs as well as low-abundance miRNAs. This powerful strategy has been applied rapidly in many plant species [10; 11; 12; 13], and the number of miRNAs increases quickly. Up to now, the total number of registered miRNAs in miRBase increased to 48, 860 in 271 species in the current release 22 ([[http://www.mirbase.org/](http://www.mirbase.org/)]([http://www.mirbase.org/](http://www.mirbase.org/)), accessed on 9 October 2021). MiRNAs regulate gene expression by repressing the translation or cleavage of targeted mRNAs [14]. Numerous reports have demonstrated that miRNAs play important roles in a range of biological and metabolic processes, including organ formation and differentiation, and response to environmental biotic and abiotic stresses [15; 16; 17; 18; 19; 20; 21; 22; 23]. Some miRNAs, such as miR164 and miR166, are involved in the regulation of leaf development [24; 25; 26]. Fruit is the main consumption organ for most vegetable crops. By using miRNA sequencing, many miRNAs have been identified potentially modulating fruit development and growth. For example, 32 conserved miRNAs were differentially expressed among developmental stages in melon, and miR393 were further validated affecting melon fruit ripening [27]. Comparing miRNA abundance in three developmental stages between wild and cultivated pepper fruits, miR159 and miR172 probably were proposed possibly affecting fruit set and fruit size [28]. Based on CRISPR/Cas9-mediated knock-out of miR164 in tomato, miR164 deficient mutant developed smaller fruit than wild type fruit mainly caused by reduced pericarp cell division and expansion [29]. Wax gourd (_Benincasa hispida_ (Thunb. [PERSON], 2n = 2x = 24) is an important vegetable crop of Cucurbitaceae family. The fruit size of wax gourd is diverse, ranging from 0.5 kg wild wax gourd to more than 20 kg cultivated wax gourd under normal cultivation conditions. Wax gourd could be stored over 6 months under natural conditions and has a long shelf-life, therefore it plays a significant role in ensuring the annual supply and regulating the off-season of vegetables [30]. Furthermore, it is also used for the treatment of many diseases or disorders [31; 32]. Recently, the release of the reference genome of wax gourd [33] facilitated fundamental research in wax gourd. However, until now, there is no report focusing on the miRNA profile of wax gourd considering their vital biological roles. In order to obtain comprehensive knowledge of miRNAs and their target genes in wax gourd, high-throughput sequencing and bioinformatic techniques were employed to acquire the first inventory of wax gourd miRNAs population.