Isolation of female-specific AFLP markers and molecular identification of genetic sex in half-smooth tongue sole (Cynoglossus semilaevis).
The sex-specific molecular marker is a useful gene resource for studying sex- determining mechanisms and controlling fish sex. Artificially produced male and female half-smooth tongue sole (Cynoglossus semilaevis) were used to screen sex-specific amplified fragment length polymorphism (AFLPs) molecular markers. The phenotypic sex of 28 tongue soles was determined by histological sectioning of gonads. The AFLP analysis of 15 females and 13 males via 64 primer combinations produced a total of 4681 scorable bands, of which 42.11% and 43.39% of bands were polymorphic in females and males, respectively. Seven female-specific AFLP markers were identified and designated as CseF382, CseF575, CseF783, CseF464, CseF136, CseF618, and CseF305, respectively. One female-specific AFLP marker (CseF382) was amplified, recovered from the gels, cloned, and sequenced (accession no. DQ487760). This female-specific AFLP marker was converted into a single-locus polymerase-chain reaction (PCR) marker of a sequence-characterized amplified region (SCAR). A simple PCR method of using the specific primers was developed for identifying genetic sex of half-smooth tongue sole. PCR products demonstrated that the initial 15 females produced the female-specific band of about 350 bp, but the initial 13 male individuals failed to produce the band. We also investigated the applicability of the PCR primers in other tongue sole individuals. The same female-specific fragment of about 350 bp was found in the additional 59 female individuals, but not in the additional 58 male individuals. This AFLP-based molecular sexing technique may have great application potential in elucidation of sex determination mechanisms and sex control in half-smooth tongue sole.
Chen
Song-Lin
SL
Key Lab for Sustainable Utilization of Marine Fisheries Resources, Ministry of Agriculture, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao, 266071, China. chensl@ysfri.ac.cn
Li
Jing
J
Deng
Si-Ping
SP
Tian
Yong-Sheng
YS
Wang
Qing-Yin
QY
Zhuang
Zhi-Meng
ZM
Sha
Zhen-Xia
ZX
Xu
Jian-Yong
JY
D000818:Animals; D001483:Base Sequence; D005260:Female; D005412:Flatfishes / Q000235:genetics*; D005819:Genetic Markers / Q000235:genetics; D008297:Male; D008969:Molecular Sequence Data; D010053:Ovary / Q000166:cytology; D010641:Phenotype; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D011110:Polymorphism, Genetic / Q000235:genetics*; D012732:Sex Determination Analysis / Q000379:methods / Q000662:veterinary*; D013737:Testis / Q000166:cytology
D005819:Genetic Markers