Isolation and Characterization of Male-Specific DNA Markers in the Rock Bream Oplegnathus fasciatus

Abstract

Sex-specific DNA markers applicable were very useful for elucidating the sex-determination mechanism and sex control in fishes. In the present study, amplified fragment-length polymorphism (AFLP) approach with 144 primer combinations was employed to identify sex-specific markers in the rock bream. Four male-specific AFLP fragments were identified which were designated as Opl286, Opl237, Opl422, and Opl228. Further sequence analysis of the sex markers' genomic region revealed subtle differences between the males and females. We identified four male-specific single-nucleotide polymorphisms (SNPs) and a deletion of 8 bp in marker Opl286, six male-specific SNPs in marker Opl237, three male-specific SNPs in marker Opl422, and eight male-specific SNPs and 1 bp inversions in marker Opl228. Specific primers were designed based on the nucleotide variation in the sequences to develop a simple polymerase chain reaction method for identifying the genetic sex of rock bream. As a result, three out of the four male-specific markers were converted into SNP markers. The male-specific AFLP markers and AFLP-derived SNP markers were tested in 100 individuals collected from three locations around the coast of Zhoushan, yielding reproducible sex identification. These male-specific DNA markers are a useful tool for the identification of the sex-determining locus in rock bream and for guiding artificial breeding programs.

Chromosomal mapping of microsatellite repeats in the rock bream fish Oplegnathus fasciatus, with emphasis of their distribution in the neo-Y chromosome

“…Conventional cytogenetic analysis of this species showed that the male karyotype is composed of 2n = 47 chromosomes (3 m + 44a), while the female karyotype is composed of 2n = 48 chromosomes (2 m + 46a). This species is characterized by having a multiple X 1 X 1 X 2 X 2 /X 1 X 2 Y sex chromosome system [18,19]. To further understanding of sex chromosome differentiation in Oplegnathus fasciatus , we mapped the chromosomal distribution of different classes of microsatellite repeats in the genome of Oplegnathus fasciatus , focusing on their distribution within the sex chromosomes.…”

Single nucleotide polymorphism markers with applications in aquaculture and assessment of its impact on natural populations

“…PCR assays were developed for two of these markers. In the rock bream Oplegnathus fasciatus, sequence analysis revealed 19 male-specific SNPs, which enabled the development of 3 SNP markers (Xu et al, 2013). Mapping the sex determination locus in the Atlantic halibut (Hippoglossus hippoglossus) using sequencing was performed (Palaiokostas et al, 2013a).…”

A novel sex-linked microsatellite marker for molecular sexing in rock bream fish Oplegnathus fasciatus

“…1) and exhibited high homology of the predicted transcriptional enhancer factor TEF-1-like of fish species, such as Larimichthys crocea and Haplochromis burtoni. Given the male heterogamy in this species (Xu et al, 2013a, b), the malespecific alleles are assumed to be located on the Y chromosome (Shikano et al, 2011). Since the observed pattern was consistent in a large number of individuals, allelic variation in this locus can be used as a diagnostic tool for sex identification in immature individuals, as well as in adults when phenotypic criteria are deemed to be unreliable.…”

“…This species possesses a differentiated Y chromosome and is characterized by having a multiple X 1 X 1 X 2 X 2 /X 1 X 2 Y sex chromosome system (Xu et al, 2013a, b). The chromosomal mapping of repetitive DNAs has revealed that the male-specific Y chromosome displays a particular amount of distinct microsatellite repeats along both arms unlike autosomes, suggesting that the differentiation of sex chromosomes is associated with the accumulation of such repetitive sequences.…”