DOI QR코드

DOI QR Code

Development of a Multiplex PCR System for Microsatellite Genotyping of the Sea Cucumber Stichopus japonicus

해삼(Stichopus japonicus)의 microsatellite 유전자형 분석을 위한 multiplex PCR 시스템 개발

  • Received : 2017.11.06
  • Accepted : 2017.12.12
  • Published : 2017.12.31

Abstract

A multiplex PCR system comprising 14 microsatellite markers was developed for genotyping analysis of the sea cucumber Stichopus japonicus. A total of 286 samples were used to evaluate genetic polymorphisms and forensic parameters of the microsatellite loci. In a single PCR reaction, all 14 loci were uniformly amplified and a total of 269 alleles were identified. The AJ19024 locus had the largest number of alleles (46), and its discriminatory power and exclusion power were 0.99 and 0.76, respectively. The fewest alleles (8) were present at the Psj2575 locus, which provided the lowest discriminatory power (0.81) and exclusion power (0.20). The mean number of alleles, mean heterozygosity, mean discrimination power and mean exclusion power per locus were 19.21, 0.70, 0.93, and 0.46, respectively. The combined matching probability for the 14 loci was $9.64{\times}10^{-19}$, and the combined power of exclusion was 0.999995. Thus, the forensic parameters evaluated in the present study demonstrated the utility of our multiplex PCR system for biological tracing methods, such as individual identification and paternity testing, in the sea cucumber.

Keywords

References

  1. Arif IA, Khan HA, Shobrak M, Al Homaidan AA, Al Sadoon M, Al Farhan AH and Bahkali AH. 2010. Interpretation of electrophoretograms of seven microsatellite loci to determine the genetic diversity of the Arabian Oryx. Genet Mol Res 9, 259-265. http://dx.doi.org/10.4238/vol9-1gmr714.
  2. Bacher JW, Flanagan LA, Smalley RL, Nassif NA, Burgart LJ, Halberg RB, Megid WM and Thibodeau, SN. 2004. Development of a fluorescent multiplex assay for detection of MSI-High tumors. Dis Markers 20, 237-250. https://doi.org/10.1155/2004/136734
  3. Berg KD, Glaser CL, Thompson RE, Hamilton SR, Griffin CA and Eshleman JR. 2000. Detection of Microsatellite Instability by Fluorescence Multiplex Polymerase Chain Reaction. J Mol Diagn 2, 20-8. http://dx.doi.org/10.1016/S1525-1578(10)60611-3.
  4. Chen M, Gao L, Zhang W, You H, Sun Q and Chang Y. 2013. Identification of forty-five gene-derived polymorphic microsatellite loci for the sea cucumber, Apostichopus japonicus. J Genet 92, 31-35.
  5. Goudet J. 2001. FSTAT, a program to estimate and test gene diversities and fixation indices. Version 2.9.3. Retrieved from https://www2.unil.ch/popgen/softwares/fstat.htm on Jul 11, 2017.
  6. Guo SW and Thompson EA. 1992. Performing the exact test of Hardy-Weinberg proportion for multiple alleles. Biometrics 48, 361-372. http://dx.doi.org/10.2307/2532296.
  7. Guo SW and Thompson EA. 1992. Performing the exact test of Hardy-Weinberg proportion for multiple alleles. Biometrics 48, 361-372. http://dx.doi.org/10.2307/2532296.
  8. Hill CR, Butler JM and Vallone PM. 2009. A 26 plex autosomal STR assay to aid human identity testing. J Forensic Sci 54, 1008-1015. https://doi.org/10.1111/j.1556-4029.2009.01110.x
  9. Ivanova NV, Dewaard JR and Hebert PDN. 2006. An inexpensive, automation-friendly protocol for recovering high-quality DNA. Mol Ecol Notes 6, 998-1002. https://doi.org/10.1111/j.1471-8286.2006.01428.x
  10. Jeffreys AJ, Wilson V and Thain SL. 1985. Hypervariable minisatellite regions in human DNA. Nature. 314, 67-73. https://doi.org/10.1038/314067a0
  11. Jones DA. 1972. Blood samples: probability of discrimination. J Forensic Sci 12, 355-359. https://doi.org/10.1016/S0015-7368(72)70695-7
  12. Kang JH, Kim YK, Kim MJ, Park JY, An CM, Kim BS, Jun JC and Kim SK. 2011 Genetic differentiation among populations and color variants of sea cucumbers (Stichopus japonicus) from Korea and China. Int J Biol Sci 30, 323-32.
  13. Kanno M, Li Q and Kijima A. 2005. Isolation and characterization of twenty microsatellite loci in japanese sea cucumber (Stichopus japonicus). Mar Biotechnol 7, 179-183. http://dx.doi.org/10.1007/s10126-004-0006-3
  14. Kim IS and LEE SJ. 2016. Incheon Customs Press Release. Retrieved form http://www.customs.go.kr/kcshome/cop/bbs/selectBoard.do?bbsId=BBSMSTR_1075&layoutMenuNo=20716&nttId=1083 on May 15, 2017.
  15. Kim MJ, Choi TJ and An HS. 2008. Population genetic structure of sea cucumber, Stichopus japonicus in Korea using microsatellite markers. Aquaculture Res 39, 1038-1045. https://doi.org/10.1111/j.1365-2109.2008.01962.x
  16. Krenke BE, Tereba A, Anderson SJ, Buel E, Culhane S, Finis CJ, Tomsey CS, Zachetti JM, Masibay A, Rabbach DR, Amiott EA and Sprecher CJ. 2002. Validation of a 16-locus fluorescent multiplex system. J Forensic Sci 47, 773-777.
  17. Kruger J, Fuhrmann W, Lichte KH and Steffens C. 1968. Zur Verwendung des Polymorphismus der sauren Erythrocytenphosphatase bei der Vaterschaftsbegutachtung. Dtsch Z Gerichtl Med 64, 127-146. https://doi.org/10.1007/BF00586813.
  18. Park KJ, Ryu SO, Baek YS, Kim YS, Kang HW and Han HS. 2013. Substrate Characteristics of Sea Cucumber Stichopus japonicus Habitats in the West Coast of Korea. Korean J Fish Aquat Sci 46, 886-891. http://dx.doi.org/10.5657/KFAS.2013.0886.
  19. Paulay G. 2014. Holothuroidea. World Register of Marine Species. Retrieved from http://www.marinespecies.org/aphia.php?p=taxdetails&id=760198 on May 15, 2017.
  20. Purcell SW, Conand C, Uthicke S and Byrne M. 2016. Ecological roles of exploited sea cucumbers. Oceanography and Marine Biology. An Annual Review 54, 367-386.
  21. Tereba A. 1999. Tools for analysis of population statistics, Profiles in DNA, vol 2. Promega Corporation Press, Madison, U.S.A., 14-16.
  22. Turrina S, Ferrian M, Caratti S and De Leo D. 2014. Evaluation of genetic parameters of 22 autosomal STR loci (PowerPlex(R) Fusion System) in a population sample from Northern Italy. Int J Legal Med 128, 281-283. http://dx.doi.org/10.1007/s00414-013-0934-4.
  23. Walsh PS, Fildes, NJ and Reynolds R. 1996. Sequence analysis and characterization of stutter products at the tetranucleotide repeat locus vWA. Nucleic Acids Res 24, 2807-2812. https://doi.org/10.1093/nar/24.14.2807
  24. Zhan AB, Bao ZM, Lu W, Hu XL, Peng W, Wang M and Hu J. 2007. Development and characterization of 45 novel microsatellite markers for sea cucumber (Apostichopus japonicus). Mol Ecol Notes 7, 1345-1348. https://doi.org/10.1111/j.1471-8286.2007.01876.x