Candidate gene identification of ovulation-inducing genes by RNA sequencing with an in vivo assay in zebrafish

PLoS One. 2018 May 1;13(5):e0196544. doi: 10.1371/journal.pone.0196544. eCollection 2018.

Abstract

We previously reported the microarray-based selection of three ovulation-related genes in zebrafish. We used a different selection method in this study, RNA sequencing analysis. An additional eight up-regulated candidates were found as specifically up-regulated genes in ovulation-induced samples. Changes in gene expression were confirmed by qPCR analysis. Furthermore, up-regulation prior to ovulation during natural spawning was verified in samples from natural pairing. Gene knock-out zebrafish strains of one of the candidates, the starmaker gene (stm), were established by CRISPR genome editing techniques. Unexpectedly, homozygous mutants were fertile and could spawn eggs. However, a high percentage of unfertilized eggs and abnormal embryos were produced from these homozygous females. The results suggest that the stm gene is necessary for fertilization. In this study, we selected additional ovulation-inducing candidate genes, and a novel function of the stm gene was investigated.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Female
  • Gene Editing
  • Gene Knockout Techniques
  • Genomics
  • Ovulation / genetics*
  • Phenotype
  • Sequence Analysis, RNA*
  • Zebrafish / genetics*
  • Zebrafish / physiology*
  • Zebrafish Proteins / chemistry
  • Zebrafish Proteins / deficiency
  • Zebrafish Proteins / genetics

Substances

  • Zebrafish Proteins
  • stm protein, zebrafish

Grants and funding

This work was supported by the Grants-in-Aid for Scientific Research on Priority Areas from the Ministry of Education, Culture, Sports, Science and Technology of Japan and JSPS KAKENHI Grant Number 25440154 and 16K07419 (to TT). This work was also supported by MEXT KAKENHI (No. 221S0002). The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.