Identification of differentially expressed microRNAs from a male sterile Ponkan mandarin (Citrus reticulata Blanco) and its fertile wild type by small RNA and degradome sequencing

Abstract

Male sterility, which could result from abnormal anther development, is a main contributor to seedless fruit production in higher plants, including citrus. MicroRNAs (miRNAs) play important roles in plant development by regulating various biological processes. In this study, by high-throughput sequencing of small RNA libraries constructed from two Ponkan mandarin (Citrus reticulata Blanco) lines, i.e., male fertile and seedy “Egan No. 1” and its male sterile and seedless mutant “Qianyang seedless,” a total of 156 known miRNAs from 32 families and 24 novel miRNAs were identified. Analyses of the expression profiles in sequencing data showed that 71 known miRNAs and 11 novel miRNAs were differentially expressed between the seedy and seedless types. By degradome sequencing, a total of 138 targets were identified for 44 miRNA families, among which 5 targets were experimentally validated by RLM-5′RACE. Consistent with the results in previous studies, target annotation demonstrated that most targets of the known miRNAs were transcription factors. Forty-one targets were also detected to be cleaved by 16 novel miRNAs. Expression profile analyses by qRT-PCR revealed a negative regulation between eight known miRNA and their targets during anther development. Our research provides new information about the miRNA regulatory network during anther development and further advances the study about the mechanism of citrus male sterility.