- http / /xuebao. jxau. edu. cn Acta Agriculturae Universitatis Jiangxiensis E - mail ndxb7775@ sina. com mtdna Cytb *.. / Coilia nasus DNA b F DNA b bp h. 9 S 9 9 π. K. 7. b S95. 7 A - - - Studies on the Genetic Diversity of Farmed and Wild Populations of Coilia nasus by Analysing Mitochondrial DNA Cyt b genes WEI Guang-lian XU Gang-chun GU Ruo-bo LI Jian-lin XU Pao *. College of Fisheries Nanjing Agriculture University Wuxi China. Key Open Laboratory of Genetic Breeding and Aquaculture Biology of Freshwater Fishes Ministry of Agriculture Freshwater Fisheries Research Center Chinese Academy of Fishery Sciences Wuxi China Abstract To provide a system analysis of differences in genetic diversity and scientific evidences of conservation of the germ plasm resources and industrialized artifical culture of Coilia nasus the genetic diversity and genetic structure of Coilia nasus were assessed from three populations by using mitochondrial Cyt b sequences. The complete mitochondrial Cyt b gene sequence is bp. Twelve haplotypes were detected from the individuals and the haplotype diversity h was. 9. And variation sites were defined of which 9 were singleton polymorphic sites and the other 9 were parsimony informative polymorphic sites. The nucleotide diversity π was. the average nucleotide difference K was. 7 and the average genetic distance was.. The result showed that genetic parameters and heredity diversities of farmed Coilia nasus - - - 7 - BADB5 JBFA7 5 DZ- BE 97 E - mail w59577@. com * E - mail xup@ ffrc. cn
mtdna Cytb 7 and wild Coilia nasus were higher than those of Coilia nasus taihuensis which meant farmed Coilia nasus populations still had a high genetic diversity. Key words Coilia nasus farmed population wild population Cyt b genetic diversity Coilia nasus Clupeiformes Engraulidae Coilia - 7-5 7-9 - - - b cytochrome b Cyt b mtdna 7-9 b.. ~. g. 9 g. ~. g. g 7 9. ~. g. g eppendorf -. DNA. 5 ml eppendorf 7 μl SET. 5 μl % SDS μl mg /ml K 55 / DNA TE DNA DNA DNA DNA 5 ng /μl. PCR L7 H595 Cyt b L7 5'-GACTT- GAAAAACCACCGTTG-' H595 5'-CTCCGATCTCCGGATTACAAGAC-' 5 μl DNA 5 ~ ng reaction buffer. 5 μl μmol /L dntp. μmol / L Taq U PCR 9 min 9 s 5 min 7 min 5 7 min PCR g /L 5'-GCGGATTCTCCGTAGACAAC-' 5'-GTAAAATTGTCTGGGTCTCC-'
. Clustal W. DnaSP version. S h π K MEGA version. Kimura NJ Neighbor - joining Bootstrap. Cyt b PCR bp PCR GenBank mtdna Cyt b NC _ 9579 Clastal X 99% M DL mtdna Cyt b Cyt b PCR Fig. Electrophoresis map of Cyt b gene PCR products farmed C. nasus Cyt b bp b A C G T. % 7. 9%. % 7. % A T 5. 5% C + G. 5% b A C G T G Population Cyt b Tab. The Nucleotide Frequencies of Cyt b gene from three populations % Base A G C T migratory C. nasus. 7. 5 7. 7. 7 farmed C. nasus.. 7. 5. C. nasus taihuensis. 7. 5 7. 7.. S. 5% 9 9 h. 9 π. K. 7 Kumar. ~. 9 7. Cyt b Kumar. Engraulis japonicus Engraulis encrasicolus mtdna Cytb HM_9 NC_9579 MEGA. bootstrap analysis BP
mtdna Cytb 9 Tab. DNA Cyt b Variation sites of Cyt b gene of mitochondrial DNA from three populations Variationsite 5 9 7 5 5 7 9 9 7 9579 - C C G G A T C G G A G A G A T T T G cy9 T... G. T A.. A. T G.... cy T... G. T...... G.... cy7 T... G. T...... G.... cy T... G. T...... G.... cy7 T... G. T...... G.... cy T.. A G. T A.. A.. G.... cy T.. A G C T A.. A.. G.... cy T T.. G. T...... G... T cy T... G. T. T.... G.... cy T.. A G. T A..... G.... cy5 T.. A G. T A..... G.... hj..... C............ hj..... C... G........ hj.................. hj5..... C... G........ hj7.. A............... hj..... C... G........ hj9..... C............ ys7..... C............ ys........... T...... ys..... C............ ys..... C........ A A A. cy hj ys cy farmed C. nasus hj C. nasus taihuensis ys migratory C. nasus. Tab. Parameter Populational genetic diversity in three populations Migratory C. nasus Farmed C. nasus C. nasus taihuensis Numberofsamples n 7 Numberofhaplotype h / Numberofvariablesites S singleton polymorphic sites / parsimony informative polymorphic sites. 5 5 / 5 / / Haplotypediversity Hd... Nucleotidediversity π... Average nucleotide difference K. 5. 59. 59 Kumar Kumar genetic distance. ~.. ~.. ~.
Tab. The average genetic distance among three populations DNA DNA Population Farmed C. nasus C. nasus taihuensis C. nasus taihuensis. Migratory C. nasus. 5. 7 Cyt b DNA Cyt b b bp S. 5% π cy hj ys fj cy farmed C. nasus hj C. nasus taihuensis ys migratory C. nasus fj Coilia mystus. 5 Cyt b Fig. Molecular phylogenetic tree based on the Cyt b sequences construced by NJ method π... Founder effect F F 9 7 7 Billington % Cyt b ~. % % 5 9. 5%. 7%. %
mtdna Cytb - DNA DNA RAPD RFLP SSR Whitehead P J P. FAO species catalogue M. Rome United Nations Development Programmer 95 7-7.. J. 5 9-9. Shi D Gong D. Strategies of resource conservation for Coilia ectenes in the Yangtze Estuary J. Marine Science 5 9-97. Li Y X Xie S G Li Z J et al. Gonad development of an anadromous fish Coilia ectenes Engraulidae in lower reach of Yangtze River China J. Fisheries Science 7 7 -. 5. J. 7.. J. 5 -. 7. J. 9-5. Haacke C Hess M Melzer R R et al. Fine structure and development of the retina of the grenadier anchovy Coilia nasus Engraulididae Clupeiformes J. Journal of Morphology - 55. 9. J. 5 5 -.. J. 977 9 -. Yang J Arai T Liu H et al. Reconstructing habitat use of Coilia mystus and Coilia ectenes of the Yangtze River estuary and of Coilia ectenes of Taihu Lake based on otolith strontium and calcium J. Journal of Fish Biology 9-5.. J. 99 9 9-5.. J. 77-777. Cheng Q Q Lu D R Ma L. Morphological differences between close populations discernible by multivariate analysis a case study of genus Coilia Teleostei Clupeiforms J. Aquatic Living Resources 5 7-9. 5 Cheng Q Q Lu D R. PCR - RFLP analysis of cytochrome b gene does not support Coilia ectenes taihuensis being a subspecies of Coilia ectenes J. Journal of Genetics 5 7 -.. b J. 5 -. 7 Bennetts R Q Grady J M Rohde F C et al. Discordant patterns of morphological and molecular change in broadtail madtoms genus Noturus J. Mol Ecol 999 5-59. Mcveigh H P Davidson W S. A salmonid phylogeny inferred from mitochondrail cytochrome b gene sequences J. Fish Biol 99 9 suppl 77 -. 9 Cantatore P Roberti M Pesole G. Evolutionary analysis of cytochrome b sequence in some perciformes evidence for a slower rate of evolution than in mammals J. Mol Evol 99 9 59-597. Xiao W H Zhang Y P Liu H Z. Molecular systematics of xenocyprinae Teleostei Cyprinidae taxono - my biogeography and coevolution of a special group restricted in East Asia J. Molecular Phylogenetics and Evolution - 7. Thompson J D Gibson T J Pewwniak F et al. The clustal x windows interface flexible strategies for multiple sequence alignment aided by quality analysis tools J. Nucleic Acids Research 997 5 7 -. Rozas J Srnehez - DelBarrio J C Messeguer X et al. DnaSP DNA polymerism analyses by the coalescent and other methods J. Bioinformatics 9 9-97. Kumar S Tamura K Nei M. MEGA Integrated software for molecular evolutionary genetics analysis and sequence alignment J. Brief Bioinform 5 5 -.
. J. 7 5-7.. J. 7-5.. J. 7 -.. D.. 5. J. 5 -.. D. 9. 7.. J. 7 9 7-7.. J. 9-7. 9 Lucey J A. Formation and physical properties of milk protein gels J. Journal of Dairy Science 5-9. 櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗櫗 Iwin D W Kocher T D Wilson A C. Evolution of the cytochrome b gene of mammals J. Mol Evo 99 -. 5. J. - 7.. DNA RAPD J. 5 5 -. 7. J. 7 5 -. Billing ton N Hebert P D N. Mitochondrial DNA diversity in fishes and its implications for introductions J. Can J Fish Aquat Sci 99-9. 9 Stephen J O Brien Ernst Mayr. Bureaucratic mischief recognizing endangered species and subspecies J. Science 99 5 7 -.. Coilia nasus J. 9 -.. J. 57-5.