Failure of PCR-RAPD technique to differentiate sex in Mahisefied (Rutilus frisii kutum) from the South Caspian Sea
In order to identify the sex marker in Mahisefied, Rutilus frisii kutum, samples from 5 male and 5 female fish were collected from the south Caspian Sea. Polymerase chain reaction random amplified polymorphic DNA (PCR-RAPD) was performed using 124 primer sets. All bands were numbered using 1 and 0 scores corresponding to the presence or absence of bands, respectively and data were analyzed using RAPDPLOT program. Results indicated that 44 sets of primers did not show any flanking site and produced no bands, while the remaining 80 produced sharp and visible bands on polyacrylamid gel. In total, 1600 bands were scored. However, none of the bands corresponded to either the male or female fish. According to the results it has been concluded that RAPD technique failed to detect sex and cannot be considered as a robust molecular tool for sex differentiation in the studied fish. The reason may be the absence of sex chromosomes in this species or that the genes corresponding to sex differentiation are spread on different autosomal chromosomes with interaction of some environmental factors. REFERENCES Abdolmaleki, Sh. and Ghani Nejad, D. (2007). Releasing Mahi Sefied Fry and Its Role in Reconstructing the Stock of this Fish in the Iranian Coasts of the Caspian Sea, Monthly Magazine of Aquatics, 8th year, pp. 8-14 (In Persian).Allendorf, F.W ; Gellman, W.A. and Thorgaard, G.H. , 1994. Sex-linkage of two enzyme loci Oncorhynchus mykiss (rainbow trout). Heredity 72: 498- 507.Azari Takami, Gh. (1984). Principles of Fish Breeding and Culture, Organization of Aquatics Breeding and Development, Deputy Aquatic Fisheries Publications, p. 152 (In Persian).Bardakci, F., 2000. The use of random amplified polymorphic DNA (RAPD) markers in sex discrimination in Nile Tilapia, Oreochromis niloticus (Pisces: Cichlidae), Turk J Biol, pp. 169-175.Baroiller J. F. and D'cotta, H., 2001. Environment and sex determination in farmed fish. Comp Biochem Physiol C Toxicol Pharmacol 130: 339-409.Callejas, C. and Ochando, M.D., 2002. Phylogenetic relationships among Spanish Barbus Species(Pisces, Cyprinidae) shown by RAPD markers. Heredity 89: 36-43.Callejas. C. and Ochando, M.D., 2001. Molecular identification (RAPD) of the eight species of the genus Barbus (Cyprinidae) in the lberian peninsula. Jornal of Fish Biology. Volume 59: 1589-1599.Congiu, L.; Rossi, R. and Colom. G., 2002. Population analysis of the sand smelt Atherina boyeri (teleostei. Atherinidae), from Italian coastal lagoons by random amplified polymorphic DNA. Marine Ecology, 1, 229: 279-289.Cui, J. Z.; Shen, X. Y.; Gong, Q. L.; Yang, G.P. and Gu, Q. Q., 2006. Identification of Sex markers by cDNA-AFLP in (Takifugu rubripes). Aquaculture, 25: 30-36.Devlin, R. H. and Nagahama, Y., 2002. Sex determination and sex differentiation in fish: an overview of genetic, physiological, and environmental influences. Aquaculture 208, pp. 191- 364.Ellegren, H., 2001. Hens, cocks and avian sex determination. A quest for genes on Zor W? Embo reports, 2: 192- 196.Fernando, A. A. and Phang, V. P. E., 1989. Inheritance of the tuxedo and blond tuxedo color pattern phenotypes of the guppy, Poecilia reticulate. Proceeding of the Second Asian Fisheries Forum, Tokyo, Japan, the Second Asian Fisheries Forum, Tokyo, Japan, pp. 487- 490.Froschauer, A., Kortin, C., Aoki, T. and Asakawa, S. , 2005. Construction and initial analysis of bacteria artificial chromosome (BAC) Cotigs from the sex determination region of the platyfish (Xiphophoorus maculatus). Gene, 295: 247-254.Griffiths, R.; Orr, K.J., Adam, A. and Barber, I., 2000. DNA sex identification in the theree spined stickleback. J. Fish Biol., 57: 1331- 1334.Horng, Y.M.; Wu, C., Wang, Y.C. and Huang, MC, 2006. A novel molecular genetic marker for gender determination of pigeons, Theriogenology 65: 1759 - 1768.Itrurra, P., Medrano, J.F., Bagley, M., Lam, N., Vergara, N. and Marin, JC. , 1998. Identification of sex chromosome molecular, marker using RAPDs and fluorescent in situ hybridization in rainbow trout. Genetica, 101: 209- 213.Iturra, P., Lam, N., Fuente, M., Vergara, N. and Medrano, J.F., 2001. Characterization of sex chromosomes in rainbow trout and coho salmon using Fluorescent In Situ Hybridization (FISH). Genetica 111, pp. 125 -131.Jaillon, O., Aury, J.M., Petit, J. and Stange-thomann, N., 2004. Genome duplication in the teloset fish Tetradon nigroviridis reveals the early vertebrate proto- karyotype. Nature, 431: 946 -957.Jian zhou, C.; Xue, Y. S.; Qing, L.G.; Guan Ping, and Y. Qin. Qun, G., 2006. Identification of sex markers by cDNA-AFLP in (Takifugu rubripes). Aquaculture, 257: 30-36.Keyvanshoukouh, S; Pourkazemi M. and Kalbasi M.R. (2004). Investigating the Possibility of Determining the Sex of Huso huso Using the PCR-RAPD Method, Iranian Fisheries Scientific Magazine, No.1, 13th year, Spring 2004, pp. 149-162 (In Persian).Kovacs, B., Egedi, S.; Bartfai, R. and Orban, L., 2001. Male-specific DNA markers from African catfish (Clarias gariepinus). Genetica, 110: 267-276.Levin, I.; Crittenden, L.B. and Dodgson, J.B., 1993. Genetic map of the chicken. Z chromosome using random amplification polymorphic DNA. Genomics 16: 224-230.Li, Y. ; Hill, J.A. ; Yue, G.H. ; Chen, F. and Orban, L., 2002. Extensive search does not identify genomic sex markers in Tetradon nigroviridis. Journal of Fish Biology. 61: 1314-1317.MaGowan, C. and Davidson, W.S., 1998. The RAPD technique fails to detect a male-specific genetic marker in Atlantic salmon. Journal of Fish Biology, 53: 1134- 1136.Martinez, E. A.; Destombe, C.; Quillet, M. C. and Valero, M., 1999. Identification of random amplified polymorphic DNA (RAPD) markers highly linked to sex determination in the red alga Gracilaria gracilis. Molecular Ecology, 8: 1533-1538.Matsuda, M.; Matsuda, C.; Hamaguchi, S. and Sakaizumi, M., 1998. Identification of the sex chromosomes of the medaka, Oryzias latipes, by fluorescence in situ hybridization. Cytogenet. Cell Genet. 82: 257- 262.McQuown, E.; Sloss, B. L.; Sheehan, R. J. and May, B., 2000. Microsatellite analysis of genetic variation in Sturgeon (Acipenseridae): new primer sequence for Scaphirhychus and Acipenser. Trans. Am. Fish. Soc. 129: 1380-1388.Penalva, L. and Sanchez, L., 2003. RNA binding protein sex- lethal (sxl) and control of Drosophila sex determination and dosage compensation. Microbial Mol. Biol Rev, 67, pp. 343-359.Pourkazemi, M. , 1996. Molecular and biochemical genetic analysis of sturgeon stocks from the South Caspian Sea. PhD Dissertation, University of Wales, Swansea. 260 p.Ravelo, C.; Magarinos, S. and Toranzo, A.E., 2002. Molecular fingerprinting of fish pathogenic Lactoccus garvieae strains by random amplified polymorphic DNA analysis. Jornal of Clinical Microbiology, pp. 751-756.Razavi Sayyad B.A. (1990). Evaluation and Management of Economic Fish Stock of the Caspian Sea, Fishery Research organization, Guilan Province, Bandar Anzali, p. 86 (In Persian).Roest Crollius, H. and Weissebach, J., 2005. Fish genomic and biology. Genom Res 15, pp. 1675-1682.Schartl, M., 2004. Sex chromosome evolution in non mammalian vertebrates. Curr Opin Genet Dev. 14: 634-641.Sun, X. Liang, L., 2003. A genetic linkage map of common carp (Cyprinus carpio L.) and mapping of a locus associated with cold tolerance. Aquaculture, 238: 165-172. Tave, D. 1993. Genetics for fish hatchery managers 2nd edn. Van Nostrand Reinhold, New York, 415 p.Wardell, B.; Sudweeks, J. D.; Meeker, N. D.; Estes, S. S.; Woodward, S.R.; and Teuscher, C., 1993. The identification of Y chromosome-linked markers with arbitrary primers. Nucleic Acids Reserch, 18: 7213-7218. Welsh, J. and McClelland, M., (1990). Fingerinting genomes using PCR with arbitrary primers. Nucleic Acids Research, 18: 7213-7218.Williams, J. G. K., Kabelik, A.R., Liva, K. J., Rafalski, J.A., and Tingey, V. , (1990). DNA polymorphism Amplified by arbitrary primers are useful as genetic markers. Nucleic Acid Research, 18: 6531-6535.Würtz, S, Gaillard, S, Barbisan, F, Carle, S, Congiu, L., Forlani, A, Aubert, J, Kirschbaum, F, Tosi, E, Zane, L. and Grillasca, J (2006). Extensive screening of sturgeon genomes by random screening techniques revealed no sex-specific marker. Aquaculture, 258: 685-688.