Screening and identification of male‐specific DNA fragments in common carps Cyprinus carpio using suppression subtractive hybridization

Abstract

In this study, a sex subtractive genomic DNA library was constructed using suppression subtractive hybridization (SSH) between male and female Cyprinus carpio. Twenty-two clones with distinguishable hybridization signals were selected and sequenced. The specific primers were designed based on the sequence data. Those primers were then used to amplify the sex-specific fragments from the genomic DNA of male and female carp. The amplified fragments from two clones showed specificity to males but not to females, which were named as Ccmf2 [387 base pairs (bp)] and Ccmf3 (183 bp), respectively. The sex-specific pattern was analysed in a total of 40 individuals from three other different C. carpio. stocks and grass carp Ctenopharyngodon idella using Ccmf2 and Ccmf3 as dot-blotting probes. The results revealed that the molecular diversity exists on the Y chromosome of C. carpio. No hybridization signals, however, were detected from individuals of C. idella, suggesting that the two sequences are specific to C. carpio. No significant homologous sequences of Ccmf2 and Ccmf3 were found in GenBank. Therefore, it was interpreted that the results as that Ccmf2 and Ccmf3 are two novel male-specific sequences; and both fragments could be used as markers to rapidly and accurately identify the genetic sex of part of C. carpio. This may provide a very efficient selective tool for practically breeding monosex female populations in aquacultural production.

Zebrafish sex: a complicated affair

“…Polymerase chain reaction (PCR)-based methods such as random amplified polymorphic DNA (RAPD) [86] and amplified fragment length polymorphism (AFLP) [87] are the most popular molecular methods for isolation of sex markers, but other procedures have also been used [88]. Information obtained from sex markers can be used to identify the type of GSD mechanism the organism is utilizing (review: [89]).…”

A newly developed genetic sex marker and its application to understanding chemically induced feminisation in roach (Rutilus rutilus)

“…The best known example is DMY, a Y chromosome‐specific duplicate of an autosomal gene called dmrt1 in medaka ( Oryzias latipes ) identified through two independent approaches, namely a positional cloning strategy and a candidate gene approach (Matsuda et al., 2002; Nanda et al., 2002). In the past, various molecular methods have led to the isolation of sex‐specific markers in various fish species, including three‐spined stickleback ( Gasterosteus aculeatus ), fathead minnow ( Pimephales promelas ), half smooth tongue sole ( Cynoglossus semilaevis ), Nile tilapia ( Oreochromis niloticus ), African catfish ( Clarias gariepinus ) and common carp ( Cyprinus carpio ) (Chen, Du, Yue, Dang, & Chang, 2010; Chen et al., 2009; Chen et al., 2007; Ezaz et al., 2004; Griffiths, Orr, Adam, & Barber, 2000; Kovács, Egedi, Bartfai, & Orban, 2000; Olmstead et al., 2011). In other fish species, including the three‐spined stickleback, sex‐linked allozyme markers have been identified (Allendorf, Gellman, & Thorgaard, 1994; Liu, Goudie, Simco, & Davis, 1996; Volff et al., 2013; Withler, McPhail, & Devlin, 1986), but in these cases, sex linkage could be a consequence of sex‐specific gene expression of autosomal genes (Devlin & Nagahama, 2002).…”

Allelic segregation of sex‐linked microsatellite markers in Nile tilapia (Oreochromis niloticus) and validation of inheritance in YY population

“…Sex‐identifying molecular markers have significant implications in genetic improvement programme for many aquaculture species where male and female fish show significant differences in economically important traits (Piferrer, Ribas, & Díaz, ). Sex‐specific DNA markers have been isolated from several important aquaculture fish, such as African catfish Clarias gariepinus (Kovacs, Egedi, Bartfai, & Orban, ), half‐smooth tongue sole Cynoglossus semilaevis (Chen et al, ), common carp Cyprinus carpio (Chen, Du, Yue, Dang, & Chang, ) and rock bream Oplegnathus fasciatus (Xu, Lou, Xu, Li, & Geng, ). Among tilapias, markers linked to sex have been identified in O. niloticus , O. aureus , O. mossambicus , O. karongae , T. mariae , T. zillii and their hybrids (Campos‐Ramos, Harvey, McAndrew, & Penman, ; Cnaani, Zilberman, Tinman, Hulata, & Ron, ; Eshel et al, ; Ezaz, Myers, Powell, McAndrew, & Penman, ; Guyon et al, ; Lee et al, , ; Lee, Penman, & Kocher, ; Palaiokostas et al, , ; Shirak et al, , ) and mapped to different linkage groups (LG).…”

“…Sex-specific DNA markers have been isolated from several important aquaculture fish, such as African catfish Clarias gariepinus (Kovacs, Egedi, Bartfai, & Orban, 2000), half-smooth tongue sole Cynoglossus semilaevis (Chen et al, 2008), common carp Cyprinus carpio (Chen, Du, Yue, Dang, & Chang, 2010)…”