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. 2020 Mar 13;10(1):4626.
doi: 10.1038/s41598-020-60946-y.

Genome-wide identification of miRNAs and their targets during early somatic embryogenesis in Dimocarpus longan Lour

Xiaoping Xu  1 , Xiaohui Chen  1 , Yan Chen  1 , Qinglin Zhang  1 , Liyao Su  1 , Xu Chen  1 , Yukun Chen  1 , Zihao Zhang  1 , Yuling Lin  2 , Zhongxiong Lai  3
Affiliations

Genome-wide identification of miRNAs and their targets during early somatic embryogenesis in Dimocarpus longan Lour

Xiaoping Xu et al. Sci Rep. .

Abstract

miRNAs are endogenous regulatory factors that play pivotal roles in post-transcriptional regulation. However, their specific roles in early somatic embryogenesis (SE) remain unclear. Study of the SE system is fundamental for clarifying the molecular mechanisms in Dimocarpus longan. We identified 289 known miRNAs from 106 different miRNA families and 1087 novel miRNAs during early longan SE, including embryogenic callus (EC), incomplete pro-embryogenic culture (ICpEC), globular embryo (GE), and non-embryogenic callus (NEC). The abundances of known miRNAs were concentrated in GE. The differentially expression (DE) miRNAs showed five expression patterns during early SE. Largely miRNAs were expressed highly and specially in EC, ICpEC, and GE, respectively. Some miRNAs and putative target genes were enriched in lignin metabolism. Most potential targets were related to the pathways of plant hormone signal transduction, alternative splicing, tyrosine metabolism and sulfur metabolism in early longan SE. The regulatory relationships between dlo-miR166a-3p and DlHD-zip8, dlo-miR397a and DlLAC7, dlo-miR408-3p and DlLAC12 were confirmed by RNA ligase-mediated rapid amplification of cDNA ends. The expression patterns of eight DE miRNAs detected by qRT-PCR were consistent with RNA-seq. Finally, the miRNA regulatory network in early SE was constructed, which provided new insight into molecular mechanism of early SE in longan.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1
Figure 1
Size distributions of miRNAs in the early SE in longan.
Figure 2
Figure 2
Quantitative distribution of 24nt, 21nt miRNA in the early SE and NEC of longan. (A) represents the distribution of 24nt miRNA at different stages of longan; (B) represents the distribution of 21nt miRNA at different stages of longan.
Figure 3
Figure 3
The number of known and novel up-regulation/down-regulation of miRNA in each group in the early SE and NEC.
Figure 4
Figure 4
Differential expression pattern of miRNA in early SE and NEC of longan.
Figure 5
Figure 5
Identification of cleavage sites between dlo-miR166a-3p, dlo-miR397a and dlo-miR408-3p and their target genes inlongan. “(A) cleavage site mapping of miR166a-3p target gene. mRNA sequence of DlHD-Zip8 is aligned with dlo-miR166a-3p. (B) cleavage site mapping of dlo-miR397a target gene. mRNA sequence of DlLAC7 is aligned with dlo-miR397a. (C) cleavage site mapping of dlo-miR408-3p target gene. mRNA sequence of DlLAC12 is aligned with dlo-miR408-3p. Numbers indicate the fraction of cloned PCR products terminating at different positions.
Figure 6
Figure 6
qRT-PCR expression patterns of 11 miRNAs in the early SE in longan. (A) relative expression levels in qRT-PCR. (B) is log2 (FPKM) values in the RNA-Seq library. The colors ranging from red to green indicate high to low correlation. (C) is the relative expressions of six miRNAs corresponding to the heat map and their potential target genes. U6 snRNA was used as a reference gene to normalize miRNA and DlUBQ was used as a reference gene to normalize mRNA expression data. “ *” indicate significant difference at p-value < 0.05,“**” indicate significant difference at p-value < 0.01.
Figure 7
Figure 7
Possible regulation network draft of miRNA in the early SE of longan. a represent some miRNA may paticipate in the lignin pathway and regulate RNA polymerase II. b represent some miRNA may paticipate in Brassinosteroid biosynthesis, Spliceosome, Tyrosine metabolism regulate the development of early SE. c represent some DE miRNA paticipate in different early SE of longan. (Note: This image was drawn by the author, according to the summary of discussion.).
See this image and copyright information in PMC

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