Microsoft Word 吴旭东_new_.doc

Size: px

Start display at page:

Download "Microsoft Word 吴旭东_new_.doc"

堰员
7 years ago
Views:

1 38 4 Vol. 38, No ACTA HYDROBIOLOGICA SINICA Jul., doi: / Cytb 1, 2 吴旭东 2 肖伟 1, 2 连总强 3 侯玉霞 (1., ; 2., ; 3., ) : ( lanzhouensis) Cytb, ( glanis) Cytb PCR, 1138 bp Cytb 13 Cytb,, 61.38% 91.12%, 76.62% 95.52% ( meridionalis)( asotus)( cochinchinensis) Cytb, Cytb, 8.87%, / 3.21;, 14.41%, / 1.83 Cytb Cytb, 11.16%, 127, / 4.08, NJ 13 Cytb, : ; Cytb; 中图分类号 : S917 文献标识码 : A 文章编号 : (2014) ( lanzhouensis), ()(Siluridea)(),,,,, () [1], [2] ;,, [3 6] ; [7, 8] ; PCR [9] (Mitochondria), [10],, [11] b(mitochondria Cytochrome b), ATP, [12] Meyer, b, 13, [13], Cytb, [14] Cytb [15] Cytb, GenBank 收稿日期 : ; 修订日期 : 基金项目 : ( ) 通信作者 : (1967 ),,, ; amy95@163.com

2 4 : Cytb 773 Cytb, [(10±1.5) cm, (120±8) g] EcoRⅠSalⅠ T4 DNA Taq DNA ; IPTGX-gal Sigma ; (Escherichia Coli)JM109, ; pmd18-t Promga 1.2 DNA 0.5 g, [16] Spectrophotometer DNA, 50 ng/µl 1.3 Cytb (GenBank : AM ) Cytb : HD: 5-CCA AAGCGGCAAAATAAG-3; LD: 5-GGAGTTAGGG GCGGGAGT-3 PCR PCR 25 µl, 10 buffer( Mg 2+ )3 µl, dntp 2 µl, 0.5 µl, Taq 0.5 µl(2.5 U/µL), ddh 2 O 25 µlpcr : 94 5min; 94 30s, 54 30s, 72 30s, 30 ; 72 5minPCR 1%,, pmd18-t 16, JM109, LB 37 16h,, [17] 1.4 Sal I EcoR I, (2 ) DNAClub Clustal x ; Mega4.0 [18] Cytb, Cytb, /; Mega4.0 Kimura s 2 Parameter, NJ Cytb Cytb PCR PCR, 1100 bp ( 1A) T, JM109,, 2000 bp, 1100 bp ( 1B), 1 Cytb PCR Fig. 1 The whole mitochondria Cytb PCR amplification product and enzyme digestion of S. lanzhouensis M DNA Marker; ; A Cytb PCR ; B DNA M is marker; Number are lanes; A is mitochondria Cytb PCR product of S. lanzhouensis; B is mitochondria Cytb enzyme digestion of pmd18-t 2.2 Cytb DNAClub,, Clustal x, 1138 bp, Mega4.0 Cytb, Cytb 1138, 379, ATG, T GenBank, HQ Cytb 13 Cytb , ( 1), TAA

3 Tab. 1 表 1 兰州鲇与其他 13 种鱼类线粒体 Cytb 基因碱基组成及密码子比较 Comparison of base composition and codons of mitochondria Cytb genes between S. lanzhouensis and other 13 species Taxonomy Genera Species Length (bp) Number of AA Initiator Terminator (A+T) (%) S. lanzhouensis ATG T 57.7 S. asotus ATG T 55.8 S. meridionalis ATG T 56.1 S. glanis S. cochinchinensis ATG T ATG T 56.0 Tetraodontiformes Takifuku T. fasciatus ATG TAA 55.0 Perciformes Cypriniformes Thunnus Cyprinus T. thynnus C. carpio ATG T ATG T 56.0 Anguilliformes Anguilla A. japonica ATG TAA 58.0 Clupeiformes Pleuronectiformes 鳀 Engraulis Paralichthys 鳀 E. japonicas P. olivaceus ATG T ATG T 55.0 Salmoniformes Salmo S. salar ATG T 55.0 Acipenseriformes Squaliformes Acipenser Squalus A. stellatus S. acanthias ATG T ATG TAA 60.0, T, Perdices, et al. Rhamdia [19] Cytb,,,,, Cytb, Cytb [20] Mega4.0 Cytb, 13 Cytb ( 2),, Cytb, 91.12% 95.52% Mega4.0 Kimura s 2 Parameter, NJ 5 Cytb, bootstrap1000 ( 2) 2.4 Cytb Mega4.0 Cytb, Cytb ( 3), Cytb, 8.87%, / 3.21;, 14.41%, / Cytb PCR, [9] (Gen- Bank : JF895472) Mega4.0

4 4 : Cytb 775 Tab. 2 Taxonomy 表 2 兰州鲇与其他 13 种鱼类线粒体 Cytb 基因序列和氨基酸序列同源性比较 Comparoson of homology of nucleotide and amino acid of mitochondria Cytb between S. lanzhouensis and other 13 species Genera Species GenBank GenBank accession Nucleotide homology (%) Amino acid homology (%) S. asotus AF S. meridionalis AF S. glanis AM S. cochinchinensis AF Takifuku T. fasciatus AJ Thunnus T. thynnus AP Cyprinus C. carpio X Anguilla A. japonica AB 鳀 Engraulis 鳀 E. japonicas AB Paralichthys P. olivaceus AB Salmo S. salar U Acipenser A. stellatus AJ Squalus S. acanthias NC Cytb Fig. 2 Phylogenetic trees derived from mitochondria Cytb nucleotide sequences of S. lanzhouensis and other thirteen species of fishes based on Neighbor-Joining method Cytb, Cytb ( 4) Cytb, 2.63%, / 4.00; 18.45%, / 2.00; Cytb 11.16%, 127, 102, 25, / 4.08 Mega4.0 Kimura s 2-Parameter Cytb Cytb ( 5 3 W-fishes ) Cytb ( 5) Cytb, ; Cytb, ; Cytb Cytb NJ ( 3), bootstrap %,,,

5 表 3 兰州鲇与四种鲇属鱼类线粒体 Cytb 基因的碱基替换分析 Tab. 3 Number and frequencies of nucleotides substitution of mitochondria Cytb genes between S. lanzhouensis and other four species Nucleotide substitution S. glanis S. meridionalis Number of base S. asotus S. cochinchinensis (TS) A G C T Total of TS (TV) A C A T G C G T Total of TV Total of nucleotides substitution Total of bases (bp) Frequencies of nucleotides substitution (%) / TS/TV 表 4 王庆容等测定的线粒体 Cytb 基因与五种鲇属鱼类线粒体 Cytb 基因之间的碱基替代分析 Tab. 4 Number and frequencies of nucleotides substitution of mitochondria Cytb genes between which was measured by Wang Qing-Rong and other five species Number of base Nucleotide substitution S. glanis S. asotus S. cochinchinensis S. meridionalis S. lanzhouensis (TS) A G C T Total of TS (TV) A C A T G C G T Total of TV Total of nucleotides substitution Total of bases (bp) Frequencies of nucleotides substitution (%) / TS/TV Tab. 5 表 5 兰州鲇等几种鲇形目鱼类基于线粒体 Cytb 基因的遗传距离 Genetic distance of S. lanzhouensis and other species based on mitochondria Cytb Species S. lanzhouensis S. lanzhouensis S. glanis S. asotus S. cochinchinensis S. meridionalis W-fishes S. glanis S. asotus S. cochinchinensis S. meridionalis W-fishes

6 4 : Cytb Cytb Cytb Cytb Fig. 3 Phylogenetic trees derived from mitochondria Cytb gene of S. lanzhouensis cloned by this paper and measured by Wang Qing-Rong et al. and other four species of based on Neighbor-Joining method 3, Avise 0.1% 5% [21], Cytb, %, 6% [22, 23] Robert, et al., Kimura s 2-Parameter [24] ; Kartavtsev, et al. Cytb (1.55±0.56)%, (5.52± 1.34)%, (10.69±1.34)% [25] Cytb Cytb 11.66%, Kimura s 2-Parameter , Avise Kartavtsev, Cytb 2.63%, Kimura s 2-Parameter ,, 2013, ; ( ) [9],, [26] ;, Cytb Cytb, ; Cytb Cytb,,, 2,, ; : ; 7 ;, Cytb, Nelson [27] [1] Wang S, Xie Y. China Species Red List (Volume II) [M]. Beijing: Higher Education Press. 2009, [,.(). :. 2009, ] [2] Cheng X L. Division of catfish fish overview of China [J]. Acta Hydrobiologica Sinica, 1977, 6(2): [.., 1977, 6(2): ] [3] Wu X D, Zhang Q, Zhao H X, et al. A new species of catfish in Ningxia lanzhouensis and it s intensive morphological description [J]. Freshwater Fisheries, 2006, 36(3): [,,,.., 2006, 36(3): 26 29] [4] Shi L N, Zhang Q, Wu X D, et al. Individual fertility of the lanzhouensis [J]. Journal of Gansu Agricultural University, 2008, 43(1): [,,,.., 2008, 43(1): 67 70] [5] Wu X D, Lian Z Q, Hou Y X, et al. Application analysis of microsatellite markers from meriaionalis on three species of siluriformes [J]. Acta Hydrobiologica Sinica, 2011, 35(4): [,,,.., 2011, 35(4): ] [6] Wu X D, Lian Z Q, Hou Y X, et al. Genetic structure

7 analyses between wild population and artificial breeding ones of lanzhouensis [J]. Freshwater Fisheries, 2011, 41(3): [,,,.., 2011, 41(3): 34 38] [7] Qiu X C, Zhao H X, Wang Y J, et al. Nutrient analysis and nutritive evaluation in muscle of catfish lanzhouensis [J]. Fisheries Science, 2008, 27(8): [,,,.., 2008, 27(8): ] [8] Qiu X C, Zhao H X, Wang Y J, et al. In vitro digestibility coefficient and enzymolysis rate of lanzhouensis to four common feed ingredients [J]. Freshwater Fisheries, 2008, 38(8): [,,,.., 2008, 38(2): 31 35] [9] Wang Q R, Xu C, Xu C R, et al. Mitochondrial genome structure of Yellow River catfish ( lanzhouensis) and phylogenetic analysis [J]. Acta Scientiarum Naturalium Universitatis Pekinensis, 2012, 48(3): [,,,.., 2012, 48(3): ] [10] Gray M W. Origin and evolution of mitochondrial DNA [J]. Annual Review of Cell Biology, 1989, 5: [11] Xiao W H, Zhang Y P. Genetics and evolution of mitocchondrial DNA in fish [J]. Acta Hydrobiologica Sinica, 2000, 24(4): [,. DNA., 2000, 24(4): ] [12] Schoepp B, Breton J, Parot P, et al. Relative orientation of the hemes of the cytochrome bc 1 complexes from Rhodobacter Sphaeroides,Rhodospirillum rubrum,and beef heart mitochondria, A linear dichroism study [J]. Biological Chemistry, 2000, 275(8): [13] Meyer A. Shortcoming of the cytochrome b gene as a molecular marker [J]. Trends in Ecology and Evolution, 1994, 9: [14] Katriina L I, Erie B T. Molecular resolution of the systematics of a problematic group of fishes (Teleostei: Osmeridae) and evidence for morphological homoplasy [J]. Molecular Phylogenetics and Evolution, 2009, 50(1): [15] Song C B, Near T J, Page L M. Phylogenetic relations among percid fishes as inferred from mitochondrial cytochrome b DNA sequence data [J]. Molecular Phylogenetics and Evolution, 1998, 10(3): [16] Wu X D, Zhang Q, Zhao H X, et al. The method of extracting and purifing genome DNA of a catfish in the Yellow River [J]. Freshwater Fisheries, 2006, 36(1): [,,,. DNA., 2006, 36(1): 19 21] [17] Sambrook J, Russell D W. Molecular Clonging: a Laboratory Manual [M]. New York: Cold Spring Harbor Laboratory Press. 2001, [18] Tamura K, Dudley J, Nei M, et al. Mega4: Molecular evolutionary genetics analysis (MEGA) software version 4.0 [J]. Molecular Biology and Evolution, 2007, (10): [19] Perdices A, Bermingham E, Montilla A, et al. Evolutionary history of the genus Rhamdia (Teleostei: Pimelodidae) in central America [J]. Molecular Phylogenetics and Evolution, 2002, 25: [20] Peng Z, He S, Zhang Y. Phylogenetic relationships of glyptostemoid fishes(: Sisoridae) inferred from mitochondrial cytochrome b gene sequence [J]. Molecular Phylogenetics and Evolution, 2005, 31(3): [21] Avise J C. Molecular Markers, Natural History and Evolution [M]. New York: Chapman and Hall. 1994, [22] Cheng Q Q, Ma C Y, Zhuang P, et al. Genetic structure and evolution characters in three populations of Coilia mystus based on cytochrome b gene segment sequence of mitochondrial DNA [J]. Journal of Fisheries of China, 2008, 32(1): 2 6 [,,,. b 3., 2008, 32(1): 2 6] [23] Yang X G, Wang Y Q, Zhou K Y, et al. The authentication of Oviductus ranae and their original animals by using molecular marker [J]. Biopharm Bull, 2002, 25(8): [24] Ward R D,Tyler S Z, Bronwyn H I, et al. DNA barcoding Australia s fish species [J]. Philosophical Transactions Royal Society B, 2005, 360: [25] Kartavtsev Y P, Lee J S. Analysis of nucleotide diversity at the cytochrome b and cytochrome oxidase 1 genes at the population, species, and genus levels [J]. Russian Journal of Genetics, 2006, 42(4): [26] Chu X L, Zheng B S, Dai D Y, et al. Fauna Sinica-Osteich-thyes [M]. Beijing: Science Press. 1999, [,,,.. :. 1999, 87 88] [27] Meng Q W, Su J X, Miao X Z. Taxonomy of Fishes [M]. Beijing: China Agriculture Press. 1995, [,,.. :. 1995, ]

8 4 : Cytb 779 THE CLONING AND SEQUENCE ANALYSIS OF THE MITOCHONDRIAL CYTOCHROME b GENE OF SILURUS LANZHOUENSIS WU Xu-Dong 1, 2, XIAO Wei 2, LIAN Zong-Qiang 1, 2 and HOU Yu-Xia 3 (1. College of Animal Science and Technology, Gansu Agricultural University, Lanzhou , China; 2. Ningxia Fisheries Research Institute, Yinchuan , China; 3. College of Science, China Agricultural University, Beijing , China) Abstract: To clone the sequence of the mitochondrial cytochrome b (Cytb) gene of lanzhouensis, we designed a specific pair of primers from the mitochondrial genome of S. glanis. The 1138 bp S. lanzhouensis Cytb gene was obtained after PCR. Then we compared the homology of the mitochondrial Cytb gene between S. lanzhouensis and 13 other species. The results showed that these species have high homology in Cytb gene. The homology value of the nucleotides was between 61.38% and 91.12%, and the value of the amino acids was between 76.62% and 95.52%. We also analyzed the nucleotides substitution of the mitochondrial Cytb gene among S. lanzhouensis, S. glanis, S. meridionalis, S. asotus, and S. cochinchinensis, and found that the lowest frequency of nucleotides substitution occurred between S. lanzhouensis and S. asotus with the value of 8.87%, and TS/TV was The highest frequency was between S. lanzhouensis and S. cochinchinensis with the value of 14.41%, and TS/TV was Qing-Rong Wang, et al. previously reported the sequence of the mitochondrial Cytb gene of S. lanzhouensis. Here we conducted the sequence difference analysis between our results and theirs. The results showed that the frequency of nucleotides substitution was 11.16%, there were 127 variable sites, TS/TV was 4.08, genetic distance was The phylogenetic tree of S. lanzhouensis and 13 other species constructed by NJ method was consistent with the traditional taxonomic results. Key words: lanzhouensis; Mitochondria Cytochrome b; Sequence analysis

Microsoft Word 曹伏君_new_.doc

Microsoft Word 曹伏君_new_.doc 45 2 Vol.45, No.2 2014 3 OCEANOLOGIA ET LIMNOLOGIA SINICA Mar., 2014 (Lethrinus erythropterus) * ( 524025) Wright (Lethrinus erythropterus),,,, ; ; ; Q954.56 doi: 10.11693/hyhz20140100011 (Lethrinus erythropterus)