通过二代测序技术开发25个小熊猫的微卫星位点
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摘要
采用Roche 454基因组测序平台在小熊猫Ailurus fulgens的基因组中进行测序拼装,得到了大量含有微卫星片段的序列。本文选择了其中467条序列设计引物,筛选出了115个微卫星位点,最终选出25个微卫星位点,重复单元包含TAT、GATA、AAACA、AAAACA等,并应用到42只小熊猫个体上进行了分型。观察到的等位基因数为3～10;观察杂合度和期望杂合度分别为0. 143～0. 929和0. 136～0. 869;多态信息含量为0. 128～0. 843,经检测所有位点均未偏离哈迪-温伯格平衡。这些微卫星位点可应用于小熊猫圈养种群的遗传管理,在未来还可进一步作为小熊猫的种群遗传学等研究的有力工具。
We developed 467 microsatellite loci from a microsatellite-enriched database for red panda( Ailurus fulgens)using the Roche 454 Genome Sequencer Platform. From this database,we isolated and characterized 115 microsatellite loci and chose 25 to genotype in 42 red pandas,with repeat motifs like TAT,GATA,AAACA,AAAACA and so on. The number of observed alleles ranged from 3 to 10. Observed and expected heterozygosities ranged from 0. 143 to 0. 929 and from 0. 136 to 0. 869,respectively. The mean polymorphic information content was 0. 128-0. 843,and the loci were not deviated from expectation of the Hardy-Weinberg equilibrium. We recommend that these markers be used in the systematic control of genetic management for A. fulgens. These markers could also be used as powerful tools applying in the population genetic research of A. fulgens in the future.
We developed 467 microsatellite loci from a microsatellite-enriched database for red panda( Ailurus fulgens)using the Roche 454 Genome Sequencer Platform. From this database,we isolated and characterized 115 microsatellite loci and chose 25 to genotype in 42 red pandas,with repeat motifs like TAT,GATA,AAACA,AAAACA and so on. The number of observed alleles ranged from 3 to 10. Observed and expected heterozygosities ranged from 0. 143 to 0. 929 and from 0. 136 to 0. 869,respectively. The mean polymorphic information content was 0. 128-0. 843,and the loci were not deviated from expectation of the Hardy-Weinberg equilibrium. We recommend that these markers be used in the systematic control of genetic management for A. fulgens. These markers could also be used as powerful tools applying in the population genetic research of A. fulgens in the future.
引文
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Taberlet P,Griffin S,Goossens B,et al.1996.Reliable genotyping of samples with very low DNA quantities using PCR[J].Nucleic Acids Research,24(16):3189-3194.
Taberlet P,Waits LP,Luikart G.2008.Noninvasive genetic sampling:look before you leap[J].Trends in Ecology&Evolution,14(8):323-327.
Tautz D,Renz M.1984.Simple sequences are ubiquitous repetitive components of eukaryotic genomes[J].Nucleic Acids Research,12(10):4127-4138.
Zhang L,Liang X,Zhang ZH,et al.2008.Identification and characterization of ten polymorphic microsatellite loci in the red panda Ailurus fulgens[J].Conservation Genetics,9(3):787-790.
Zhang L,Zhang ZH,Shen FJ,et al.2015.Identification and characterization of polymorphic microsatellite loci in the redcrowned crane[J].Genetics&Molecular Research,14(4):15169-15176.
Zhang WP,Zhang ZH,Wei K,et al.2012.Isolation and characterization of polymorphic tri-and tetra-nucleotide microsatellite loci for the south China tiger[J].Journal of Natural History,40(38-40):2259-2263.
Zhang ZH,Shen FJ,Sun S,et al.2003.Paternity assignment of giant panda by microsatellite genotyping[J].Hereditas,25(5):504.
李明,魏辅文,冯祚建,等.2001.保护生物学一新分支学科---保护遗传学[J].四川动物,20(1):16-19.
Armstrong DP,Seddon PJ.2008.Directions in reintroduction biology[J].Trends in Ecology&Evolution,23(1):20-25.
Blouin SF,Blouin M.1988.Inbreeding avoidance behaviors[J].Trends in Ecology&Evolution,3(9):230-233.
Botstein D,White RL,Skolnick M,et al.1980.Construction of a genetic linkage map in man using restriction fragment length polymorphisms[J].American Journal of Human Genetics,32(3):314-331.
Edwards A,Civitello A,Hammond HA,et al.1991.DNA typing and genetic mapping with trimeric and tetrameric tandem repeats[J].American Journal of Human Genetics,49(4):746-756.
Estoup A,Angers B.1998.Microsatellites and minisatellites for molecular ecology:theoretical and empirical considerations[J].Advances in Molecular Ecology Nato Sciences,38(1):69-75.
Glatston A,Wei F,Than Z,et al.2015.Ailurus fulgens[DB/OL].(2018-05-10).The IUCN Red List of Threatened Species 2015:e.T714A110023718.
Jamieson IG.2011.Founder effects,inbreeding,and loss of genetic diversity in four avian reintroduction programs[J].Conservation Biology,25(1):115-123.
Kashi Y,King D,Soller M.1997.Simple sequence repeats as a source of quantitative genetic variation[J].Trends in Genetics,13(2):74-78.
Keller LF,Waller DM.2002.Inbreeding effects in wild populations[J].Trends in Ecology&Evolution,17(5):230-241.
Liang X,Zhang ZH,Zhang L,et al.2010.Isolation and characterization of 16 tetranucleotide microsatellite loci in the red panda(Ailurus fulgens)[J].Molecular Ecology Resources,7(6):1012-1014.
Margulies M,Egholm M,Altman WE,et al.2005.Genome sequencing in microfabricated high-density picolitre reactors[J].Nature,437(7056):376-380.
Marshall TC,Slate J,Kruuk LEB,et al.2010.Statistical confidence for likelihood-based paternity inference in natural populations[J].Molecular Ecology,7(5):639-655.
Meffe GK,Carroll CR.1994.The design of conservation reserves[M]//Meffe GK,Carroll CR.Principles of conservation biology.Sunderland,MA:Sinauer Associates,Inc.:265-304.
Nagy S,Poczai P,Cernák I,et al.2012.PICcalc:an online program to calculate polymorphic information content for molecular genetic studies[J].Biochemical Genetics,50(9-10):670-672.
Omote K,Nishida C,Takenaka T,et al.2012.Temporal changes of genetic population structure and diversity in the endangered Blakiston's fish owl(Bubo blakistoni)on Hokkaido Island,Japan,revealed by microsatellite analysis[J].Zoological Science,29(5):299-304.
Ryman N,Leimar O.2008.Effect of mutation on genetic differentiation among nonequilibrium populations[J].Evolution,62(9):2250-2259.
Raymond M,Rousset F.2003.A population genetic software for exact test and ecumenicism:GENEPOP,version 3.4[J].Heredity,68:248-249.
Rozen S,Skaletsky H.2000.Primer3 on the WWW for general users and for biologist programmers[J].Methods in Molecular Biology,132(3):365-386.
Selkoe KA,Toonen RJ.2006.Microsatellites for ecologists:a practical guide to using and evaluating microsatellite markers[J].Ecology Letters,9(5):615-629.
Taberlet P,Griffin S,Goossens B,et al.1996.Reliable genotyping of samples with very low DNA quantities using PCR[J].Nucleic Acids Research,24(16):3189-3194.
Taberlet P,Waits LP,Luikart G.2008.Noninvasive genetic sampling:look before you leap[J].Trends in Ecology&Evolution,14(8):323-327.
Tautz D,Renz M.1984.Simple sequences are ubiquitous repetitive components of eukaryotic genomes[J].Nucleic Acids Research,12(10):4127-4138.
Zhang L,Liang X,Zhang ZH,et al.2008.Identification and characterization of ten polymorphic microsatellite loci in the red panda Ailurus fulgens[J].Conservation Genetics,9(3):787-790.
Zhang L,Zhang ZH,Shen FJ,et al.2015.Identification and characterization of polymorphic microsatellite loci in the redcrowned crane[J].Genetics&Molecular Research,14(4):15169-15176.
Zhang WP,Zhang ZH,Wei K,et al.2012.Isolation and characterization of polymorphic tri-and tetra-nucleotide microsatellite loci for the south China tiger[J].Journal of Natural History,40(38-40):2259-2263.
Zhang ZH,Shen FJ,Sun S,et al.2003.Paternity assignment of giant panda by microsatellite genotyping[J].Hereditas,25(5):504.