A combination of genome-wide association study screening and SNaPshot for detecting sex-related SNPs and genes in Cynoglossus semilaevis

Bo Zhang; Na Zhao; Kangkang Peng; Xiaoxu He; Chun Xiu Chen; Hao Liu; Kefeng Liu; Lei Jia; Baolong Bao

doi:10.1016/j.cbd.2020.100711

A combination of genome-wide association study screening and SNaPshot for detecting sex-related SNPs and genes in Cynoglossus semilaevis

Comp Biochem Physiol Part D Genomics Proteomics. 2020 Sep:35:100711. doi: 10.1016/j.cbd.2020.100711. Epub 2020 Jul 9.

Authors

Bo Zhang¹, Na Zhao², Kangkang Peng³, Xiaoxu He⁴, Chun Xiu Chen⁴, Hao Liu⁴, Kefeng Liu⁴, Lei Jia⁵, Baolong Bao⁶

Affiliations

¹ Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources (Shanghai Ocean University), Ministry of Education, International Research Center for Marine Biosciences at Shanghai Ocean University, Ministry of Science and Technology, National Demonstration Center for Experimental Fisheries Science Education, Shanghai Ocean University, Shanghai 201306, China; Tianjin Fisheries Research Institute, Tianjin, China.
² Tianjin Medicine Biotechnology Co, Ltd, Tianjin, China.
³ Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources (Shanghai Ocean University), Ministry of Education, International Research Center for Marine Biosciences at Shanghai Ocean University, Ministry of Science and Technology, National Demonstration Center for Experimental Fisheries Science Education, Shanghai Ocean University, Shanghai 201306, China.
⁴ Tianjin Fisheries Research Institute, Tianjin, China.
⁵ Tianjin Fisheries Research Institute, Tianjin, China. Electronic address: tianjinbohaisuo@163.com.
⁶ Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources (Shanghai Ocean University), Ministry of Education, International Research Center for Marine Biosciences at Shanghai Ocean University, Ministry of Science and Technology, National Demonstration Center for Experimental Fisheries Science Education, Shanghai Ocean University, Shanghai 201306, China. Electronic address: blbao@shou.edu.cn.

PMID: 32683285
DOI: 10.1016/j.cbd.2020.100711

Abstract

Chinese tongue sole (Cynoglossus semilaevis) males and females exhibit great differences in growth rate and appearance. The species is heterogametic (ZW/ZZ) and has sex-reversed "pseudomales" that are genetically female and physiologically male. In this study, we identified eight sex-specific single nucleotide polymorphism (SNP) markers for the sex identification of C. semilaevis by using a combination of genome-wide association study (GWAS) screening and SnaPshot validation. Candidate SNPs were screened using genotyping by sequencing to perform GWAS of the differential SNPs between the sexes of C. semilaevis. The SNP loci were amplified using a multiplex PCR system and detected via SNaPshot, which enables multiplexing of up to 30-40 SNPs in a single assay and ensures high accuracy of the results. The molecular markers detected in our study were used to successfully identify normal males and pseudomales from 45 caught and 40 cultured C. semilaevis specimens. Linkage disequilibrium analysis showed that the eight SNP loci were related to each other, with a strong linkage. Moreover, we investigated the expression of prdm6 mRNA containing a missense SNP and confirmed that the gene is differentially expressed in the gonads of the different sexes of C. semilaevis; the expression of prdm6 mRNA was significantly higher in the males than in the females and pseudomales. This means prdm6 may be related to sex differentiation in C. semilaevis.

Keywords: Cynoglossus semilaevis; SNP; SNaPshot; Sex marker; prdm6.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Female
Flatfishes / genetics*
Flatfishes / growth & development
Genome-Wide Association Study
Linkage Disequilibrium
Male
Polymorphism, Single Nucleotide*
RNA, Messenger / genetics
Sex Differentiation

Substances

RNA, Messenger