Molecular Biology Reports

, Volume 41, Issue 8, pp 5467–5471 | Cite as

Characterization of 40 single nucleotide polymorphism (SNP) via T m-shift assay in the mud crab (Scylla paramamosain)



In this study, single nucleotide polymorphism (SNP) were identified, confirmed and genotyped in the mud crab (Scylla paramamosain) using T m-shift assay. High quality sequences (13, 311 bp long) were obtained by re-sequencing that contained 91 SNPs, with a density of one SNP every 146 bp. Of all 91 SNPs, 40 were successfully genotyped and characterized using 30 wild specimens by T m-shift assay. The minor allele frequency per locus ranged from 0.017 to 0.500. The observed and expected heterozygosity, and polymorphism information content (PIC) ranged from 0.000 to 0.600, from 0.033 to 0.509, and from 0.033 to 0.375, respectively, with an average of 0.142, 0.239 and 0.198 per locus. Seventeen SNPs were significantly deviated from Hardy–Weinberg equilibrium. No significant linkage disequilibrium between pairs of loci was detected after sequential Bonferroni correction (P > 0.00125). Seventeen SNPs were related with known function genes. This study provided new molecular markers for investigation of population genetic diversity, construction of genetic linkage maps and molecular marker-assisted selection in this important crustacean species.


The mud crab Scylla paramamosain SNPs Transcriptome Genetic diversity 



This study was supported by the National Non-Profit Institutes (East China Sea Fisheries Research Institute) (No. 2011M05), the National Natural Science Foundation of China (No. 31001106), the Science and Technology Commission of Shanghai Municipality (No. 10JC1418600) and the National Infrastructure of Fishery Germplasm Resources.

Supplementary material

11033_2014_3420_MOESM1_ESM.docx (40 kb)
Supplementary material 1 (DOCX 39 kb)


  1. 1.
    Shen Y, Lai Q (1994) Present status of mangrove crab (Scylla serrate (Forskal)) culture in China, NAGA: the ICLARM Quarterly, January 28–29Google Scholar
  2. 2.
    Fishery Bureau of Ministry of Agriculture of China (2012) China Fisheries Yearbook. Chinese Agricultural Press, BeijingGoogle Scholar
  3. 3.
    Ma HY, Ma CY, Ma LB (2011) Population genetic diversity of mud crab (Scylla paramamosain) in Hainan Island of China based on mitochondrial DNA. Biochem Syst Ecol 39:434–440CrossRefGoogle Scholar
  4. 4.
    Ma HY, Cui HY, Ma CY, Ma LB (2012) High genetic diversity and low differentiation in mud crab (Scylla paramamosain) along the southeastern coast of China revealed by microsatellite markers. J Exp Biol 215:3120–3125CrossRefPubMedGoogle Scholar
  5. 5.
    Takano M, Barinova A, Sugaya T, Obata Y, Watanabe T, Ikeda M, Taniguchi N (2005) Isolation and characterization of microsatellite DNA markers from mangrove crab, Scylla paramamosain. Mol Ecol Notes 5:794–795CrossRefGoogle Scholar
  6. 6.
    Xu XJ, Wang GZ, Wang KJ, Li SJ (2009) Isolation and characterization of ten new polymorphic microsatellite loci in the mud crab, Scylla paramamosain. Conserv Genet 10:1877–1878CrossRefGoogle Scholar
  7. 7.
    Yao HF, Sun DQ, Wang RX, Shi G (2012) Rapid isolation and characterization of polymorphic microsatellite loci in the mud crab, Scylla paramamosain (Portunidae). Genet Mol Res 11:1503–1506CrossRefPubMedGoogle Scholar
  8. 8.
    Ma HY, Ma CY, Ma LB (2011) Identification of type I microsatellite markers associated with genes and ESTs in Scylla paramamosain. Biochem Syst Ecol 39:371–376CrossRefGoogle Scholar
  9. 9.
    Ma HY, Ma CY, Ma LB, Zhang FY (2011) Isolation and characterization of 54 polymorphic microsatellite markers in Scylla paramamosain by FIASCO approach. J World Aquacult Soc 42:591–597CrossRefGoogle Scholar
  10. 10.
    Ma HY, Ma QQ, Ma CY, Ma LB (2011) Isolation and characterization of gene-derived single nucleotide polymorphism (SNP) markers in Scylla paramamosain. Biochem Syst Ecol 39:419–424CrossRefGoogle Scholar
  11. 11.
    Li S, Wan H, Ji H, Zhou K, Yang G (2009) SNP discovery based on CATS and genotyping in the finless porpoise (Neophocaena phocaenoides). Conserv Genet 10:2013–2019CrossRefGoogle Scholar
  12. 12.
    Ciobanu DC, Bastiaansen JWM, Magrin J, Rocha JL, Jiang DH, Yu N, Geiger B, Deeb N, Rocha D, Gong H, Konghorn BP, Plastow GS, van der Steen HAM, Mileham AJ (2010) A major SNP resource for dissection of phenotypic and genetic variation in Pacific white shrimp (Litopenaeus vannamei). Anim Genet 41:39–47CrossRefPubMedGoogle Scholar
  13. 13.
    Olsen MT, Volny VH, Berube M, Dietz R, Lydersen C, Kovacs K, Dodd R, Palsbøll PJ (2011) A simple route to single-nucleotide polymorphisms in a nonmodel species: identification and characterization of SNPs in the Artic ringed seal (Pusa hispida hispida). Mol Ecol Resour 11(Suppl. 1):9–19CrossRefPubMedGoogle Scholar
  14. 14.
    Amish S, Hohenlohe PA, Painter S, Leary RF, Muhlfeld C, Allendorf F, Luikart G (2012) RAD sequencing yields a high success rate for westslope cutthroat and rainbow trout species-diagnostic SNP assays. Mol Ecol Resour 12:653–660CrossRefPubMedGoogle Scholar
  15. 15.
    Moen T, Delghand M, Wesmajervi MS, Westgaard JI, Fjalestad KT (2009) A SNP/microsatellite genetic linkage map of the Atlantic cod (Gadus morhua). Anim Genet 40:993–996CrossRefPubMedGoogle Scholar
  16. 16.
    Xia JH, Liu F, Zhu ZY, Fu JJ, Feng JB, Li JL, Yue GH (2010) A consensus linkage map of the grass carp (Ctenopharyngodon idella) based on microsatellites and SNPs. BMC Genom 11:135CrossRefGoogle Scholar
  17. 17.
    Kongchum P, Sandel E, Lutzky S, Hallerman EM, Hulata G, David L, Palti Y (2011) Association between IL-10α single nucleotide polymorphisms and resistance to cyprinid herpesvirus-3 infection in common carp (Cyprinus carpio). Aquaculture 315:417–421CrossRefGoogle Scholar
  18. 18.
    Yu H, He Y, Wang X, Zhang Q, Bao Z, Guo X (2011) Polymorphism in a serine protease inhibitor gene and its association with disease resistance in the eastern oyster (Crassostrea virginica Gmelin). Fish Shellfish Immun 30:757–762CrossRefGoogle Scholar
  19. 19.
    Jin S, Zhang X, Jia Z, Fu H, Zheng X, Sun X (2012) Genetic linkage mapping and genetic analysis of QTL related to eye cross and eye diameter in common carp (Cyprinus carpio L.) using microsatellites and SNPs. Aquaculture 358–359:176–182CrossRefGoogle Scholar
  20. 20.
    Milano I, Babbucci M, Panitz F, Ogden R, Nielsen RO, Taylor MI, Helyar SJ, Carvalho GR, Espineira M, Atanassova M, Tinti F, Maes GE, Patarnello T, Consortium F, Bargelloni L (2011) Novel tools for conservation genomics: comparing two high-throughput approaches for SNP discovery in the transcriptome of the European hake. PLOS ONE 6:e28008PubMedCentralCrossRefPubMedGoogle Scholar
  21. 21.
    Helyar SJ, Limborg MT, Bekkevold D, Babbucci M, Houdt J, Maes GE, Bargelloni L, Nielsen RO, Taylor MI, Ogden R, Cariani A, Carvalho GR, Consortium F, Panitz F (2012) SNP discovery using next generation transcriptomic sequencing in Atlantic herring (Clupea harengus). PLoS ONE 7:e42089CrossRefPubMedGoogle Scholar
  22. 22.
    Ma HY, Yang JF, Su PZ, Chen SL (2009) Genetic analysis of gynogenetic and common populations of Verasper moseri using SSR markers. Wuhan Univ J Nat Sci 14:267–273CrossRefGoogle Scholar
  23. 23.
    Wang J, Chuang K, Ahluwalia M, Patel S, Umblas N, Mirel D, Higuchi R, Germer S (2005) High-throughtput SNP genotyping by single-tube PCR with T m-shift primers. Biotechniques 39:885–893CrossRefPubMedGoogle Scholar
  24. 24.
    Rice WR (1989) Analyzing tables of statistical tests. Evolution 43:223–225CrossRefGoogle Scholar
  25. 25.
    Sobrino S, Lareu M, Brion M, Carracedo A (2004) SNP genotyping with single base extension-tag microarrays. Int Congress Ser 1261:331–333CrossRefGoogle Scholar
  26. 26.
    Tost J, Gut IG (2005) Genotyping single nucleotide polymorphisms by MALDI mass spectrometry in clinical applications. Clin Biochem 38:335–350CrossRefPubMedGoogle Scholar
  27. 27.
    Liu H, Li S, Wang Z, Ji M, Nie L, He N (2007) High-throughput SNP genotyping based on solid-phase PCR on magnetic nanoparticles with dual-color hybridization. J Biotechnol 131:217–222CrossRefPubMedGoogle Scholar
  28. 28.
    Matsunaga T, Maruyama K, Takeyama H, Katoh T (2007) High-throughput SNP detection using nano-scale engineered biomagnetite. Biosens Bioelectron 22:2315–2321CrossRefPubMedGoogle Scholar
  29. 29.
    Hayford AE, Mammel MK, Lacher DW, Brown EW (2011) Single nucleotide polymorphism (SNP)-based differentiation of Shigella isolates by pyrosequencing. Infect Genet Evol 11:1761–1768CrossRefPubMedGoogle Scholar
  30. 30.
    Hansen MH, Young S, Jørgense HH, Pascal C, Henryon M, Seeb J (2011) Assembling a dual purpose TaqMan-based panel of single-nucleotide polymorphism markers in rainbow trout and steelhead (Oncorhynchus mykiss) for association mapping and population genetics analysis. Mol Ecol Resour 11(Suppl. 1):67–70CrossRefPubMedGoogle Scholar
  31. 31.
    Bichenkova EV, Lang Z, Yu X, Rogert G, Douglas KT (2011) DNA-mounted self-assembly: new approaches for genomic analysis and SNP detection. BBA-Gene Regul Mech 1809:1–23Google Scholar
  32. 32.
    Song MY, Kim HE, Kim S, Choi IH, Lee JK (2012) SNP-based large-scale identification of allele-specific gene expression in human B cells. Gene 493:211–218CrossRefPubMedGoogle Scholar
  33. 33.
    Liu W, Li H, Bao X, He C, Li W, Shan Z (2011) The first set of EST-derived single nucleotide polymorphism markers for Japanese scallop, Patinopecten yessoensis. J World Aquacult Soc 42:456–461CrossRefGoogle Scholar
  34. 34.
    Campbell NR, Narum SR (2012) Development of 54 novel single-nucleotide polymorphism (SNP) assays for sockeye and coho salmon and assessment of available SNPs to differentiate stocks within the Columbia River. Mol Ecol Resour 11(Suppl. 1):20–30Google Scholar
  35. 35.
    Zhang L, Guo X (2010) Development and validation of single nucleotide polymorphism markers in the eastern oyster Crassostrea virginica Gmelin by mining ESTs and resequencing. Aquaculture 302:124–129CrossRefGoogle Scholar
  36. 36.
    Pante E, Rohfritsch A, Becquet V, Belkhir K, Bierne N, Garcia P (2012) SNP detection from de novo transcriptome sequencing in the bivalve Macoma balthica: marker development for evolutionary studies. PLoS ONE 7:e52302PubMedCentralCrossRefPubMedGoogle Scholar
  37. 37.
    Elfstrom CM, Gaffney PM, Smith CT, Seeb JE (2005) Characterization of 12 single nucleotide polymorphisms in weathervane scallop. Mol Ecol Notes 5:406–409CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2014

Authors and Affiliations

  1. 1.Key Laboratory of East China Sea and Oceanic Fishery Resources Exploitation, Ministry of Agriculture, East China Sea Fisheries Research InstituteChinese Academy of Fishery SciencesShanghaiChina
  2. 2.College of Fisheries and Life ScienceShanghai Ocean UniversityShanghaiChina

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