Horticultural Science & Technology (원예과학기술지)

Korean Society of Horticultural Science (한국원예학회)
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Identification of Lettuce Germplasms and Commercial Cultivars Using SSR Markers Developed from EST

EST로부터 개발된 SSR 마커를 이용한 상추 유전자원 및 유통품종의 식별

  • Hong, Jee-Hwa (Variety Testing Division, Korea Seed & Variety Service, Ministry of Agriculture, Food and Rural Affairs) ;
  • Kwon, Yong-Sham (Variety Testing Division, Korea Seed & Variety Service, Ministry of Agriculture, Food and Rural Affairs) ;
  • Choi, Keun-Jin (Variety Testing Division, Korea Seed & Variety Service, Ministry of Agriculture, Food and Rural Affairs) ;
  • Mishra, Raghvendra Kumar (Department of Molecular Biotechnology, Konkuk University) ;
  • Kim, Doo Hwan (Department of Molecular Biotechnology, Konkuk University)
  • 홍지화 (농림축산식품부 국립종자원 재배시험과) ;
  • 권용삼 (농림축산식품부 국립종자원 재배시험과) ;
  • 최근진 (농림축산식품부 국립종자원 재배시험과) ;
  • ;
  • 김두환 (건국대학교 분자생명공학과)
  • Received : 2013.04.13
  • Accepted : 2013.07.31
  • Published : 2013.12.31
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Abstract

The objective of this study was to develop simple sequence repeat (SSR) markers from expressed sequence tags (EST) of lettuce (Lactuca sativa) and identify 9 germplasms from 3 wild species of lettuce and 61 commercial cultivars using the developed EST-SSR markers. A total of 81,330 lettuce ESTs from NCBI databases were used to search for SSR and 4,229 SSR loci were identified. The highest proportion (59.12%, 2500) was represented by trinucleotide, followed by dinucleotide (29.70%, 1256) and hexanucleotide (6.62%, 280) among SSR repeat motifs. Totally 474 EST-SSR primers were developed from EST and a random set of 267 primers was used to assess the genetic diversity among 9 germplasms and 61 cultivars. Out of 267 primers, 47 EST-SSR markers showed polymorphism between 7 cultivars. Twenty-six EST-SSR markers among 47 EST-SSR markers showed high polymorphism, reproducibility, and band clearance. The relationship between 26 markers genotypes and 70 accessions was analyzed. Totally 127 polymorphic amplified fragments were obtained by 26 EST-SSR markers and two to nine SSR alleles were detected for each locus with an average of 4.88 alleles per locus. Average polymorphism information content was 0.542, ranging from 0.269 to 0.768. Genetic distance of clusters ranged from 0.05 to 0.94 between 70 accessions and dendrogram at a similarity of 0.34 gave 7 main clusters. Analysis of genetic diversity revealed by these 26 EST-SSR markers showed that the 9 germplasms and 61 commercial cultivars were discriminated by marker genotypes. These newly developed EST-SSR markers will be useful for cultivar identification and distinctness, uniformity and stability test of lettuce.

본 연구의 목적은 상추(Lactuca sativa)의 expressed sequence tag(EST)로부터 simple sequence repeat(SSR) 마커를 개발하고, 개발된 EST-SSR 마커를 이용하여 상추의 3가지 야생종의 유전자원 9점과 61개의 유통품종을 식별하는 것이다. NCBI 데이터베이스로부터 총 81,330개의 상추 EST를 대상으로 SSR을 탐색하였고, 총 4,229개의 SSR을 발견하였다. SSR의 반복 motif 중 trinucleotide(59.12%, 2,500개)가 가장 많았고, 그 다음으로 dinucleotide(29.70%, 1,256개), hexanucleotide(6.62%, 280개) 순의 분포를 나타내었다. EST로부터 총 474개의 EST-SSR primers를 개발하였고, 이 중 267개의 primer를 9점의 유전자원과 61품종에 대한 유전적 다양성 평가에 활용하였다. 267개의 마커 중 47개의 EST-SSR 마커가 7개 품종 내에서 다형성을 보였으며, 이 중 다형성 정도와 반복 재현성 및 밴드의 선명성을 고려하여 26개의 EST-SSR 마커를 선발하였다. 최종 선발된 26개의 SSR 마커를 이용하여 70개 공시재료를 분석한 결과 대립유전자 수는 총 127개였으며, 최소 2개에서 9개의 분포를 나타내었으며 마커당 평균 대립유전자 수는 4.88개를 나타내었다. PIC평균값은 0.542로 나타났으며, 0.269-0.768의 범위를 나타내었다. 70개 공시재료의 유전적 거리는 0.05-0.94로 나타났으며, 유사도 지수 0.34를 기준으로 할 때 7개의 주요 그룹으로 나누어졌다. 26개의 EST-SSR 마커를 이용한 유전적 다양성 분석 결과 9점의 유전자원과 61개의 유통품종이 마커의 유전자형에 의해 모두 식별이 되었다. 본 연구를 통해 신규 개발된 EST-SSR 마커는 상추의 품종식별과 구별성, 균일성, 안정성 검정에 유용하게 활용될 수 있을 것으로 사료된다.

Keywords

References

  1. Anderson, J.A., G.A. Churchill, J.E. Autrigue, and S.D. Tanksley. 1993. Optimizing parental selection for genetic linkage maps. Genome 36:181-186. https://doi.org/10.1139/g93-024
  2. Bredemeijer, G.M.M., R.J. Cooke, M.W. Ganal, R. Peeters, P. Isaac, Y. Noordijk, S. Rendell, J. Jackson, M.S. Roder, K. Wendehake, M. Dijcks, M. Amelaine, V. Wickaert, L. Bertrand, and B. Vosman. 2002. Construction and testing of a microsatellite database containing more than 500 tomato varieties. Theor. Appl. Genet. 105:1019-1026. https://doi.org/10.1007/s00122-002-1038-6
  3. Esselink, G.D., M.J.M. Smulders, and B. Vosman. 2003. Identification of cut rose (Rosa hybrida) and rootstock varieties using robust sequence tagged microsatellite site markers. Theor. Appl. Genet. 106:277-286.
  4. Funk, V.A., R.J. Bayer, S. Keeley, R. Chan, L. Watson, B. Gemeinholzer, E. Schilling, J.L. Panero, B.G. Baldwin, N. Garcia-Jacas, A. Susanna, and R.K. Jansen. 2005. Everywhere but Antaractica: Using a supertree to understand the diversity and distribution of the Compositae. Biol. Skr. 55:343-374.
  5. Ge, H., H. Li, Y. Liu, X. Li, and H. Chen. 2011. Characterization of novel developed expressed sequence tag (EST)-derived simple sequence repeat (SSR) markers and their application in diversity analysis of eggplant. African J. Biotechnol. 10:9023-9031.
  6. Gupta, P.K., S. Rustgi, S. Sharma, R. Singh, N. Kumar, and H.S. Balyan. 2003. Transferable EST-SSR markers for the study of polymorphism and genetic diversity in bread wheat. Mol. Genet. Genomics 270:315-323. https://doi.org/10.1007/s00438-003-0921-4
  7. Hu, J., J. Li, F. Liang, L. Liu, and S. Si. 2010. Genetic relationship of a cucumber germplasm collection revealed by newly developed EST-SSR markers. J. Genet. 89:28-32.
  8. Huang, X. and A. Madan. 1999. CAP3: A DNA sequence assembly program. Genome Res. 9:868-877. https://doi.org/10.1101/gr.9.9.868
  9. International Union for the Protection of New Varieties of Plants (UPOV). 2010. UPOV/INF/17/1 Guideline for DNA-profiling: molecular marker selection and database construction (BMT guideline). UPOV, Geneva.
  10. International Union for the Protection of New Varieties of Plants (UPOV). 2011a. BMT/13/12 The use of molecular markers for the lettuce species. UPOV, Brasilia.
  11. International Union for the Protection of New Varieties of Plants (UPOV). 2011b. UPOV/INF/18/1 Possible use of molecular markers in the examination of distinctness, uniformity, and stability (DUS). UPOV, Geneva.
  12. Kalia, R.K., M.K. Rai, S. Kalia, R. Singh, and A.K. Dhawan. 2011. Microsatellite markers: An overview of the recent progress in plants. Euphytica 177:309-334. https://doi.org/10.1007/s10681-010-0286-9
  13. Kristkova, E., I. Dolezalova, A. Lebeda, V. Vinter, and A. Novotna. 2008. Description of morphological characters of lettuce (Lactuca sativa L.) genetic resources. Hort. Sci. (Prague) 38:113-129.
  14. Lebeda, A., E.J. Ryder, B. Grube, I. Dolezalova, and E. Kristkova. 2007. Lettuce (Asteraceae; Lactuca spp.), p. 377-472. In: R.J. Singh (ed.). Genetic resources, chromosome engineering, and crop improvement. CRC Press, Boca Raton, FL.
  15. Leigh, F., V. Lea, J. Law, P. Wolters, W. Powell, and P. Donini. 2003. Assessment of EST- and genomic microsatellite markers for variety discrimination and genetic diversity studies in wheat. Euphytica 133:359-366. https://doi.org/10.1023/A:1025778227751
  16. Rauscher, G. and I. Simko. 2013. Development of genomic SSR markers for fingerprinting lettuce (Lactuca sativa L.) cultivars and mapping genes. BMC Plant Biol. 13:11. https://doi.org/10.1186/1471-2229-13-11
  17. Reid, A., L. Hof, G. Felix, B. Rucker, S. Tams, E. Milczynska, D. Esselink, G. Uenk, B. Vosman, and A. Weitz. 2011. Construction of an integrated microsatellite and key morphological characteristic database of potato varieties on the EU common catalogue. Euphytica 182:239-249. https://doi.org/10.1007/s10681-011-0462-6
  18. Rohlf, F.J. 2000. NTSYSpc: Numerical taxonomy and multivariate analysis system, ver. 2.10b. Applied Biostatistics Inc., New York.
  19. Rozen, S. and H.J. Skaletsky. 2000. Primer3 on the WWW for general users and for biologist programmers, p. 365-386. In: S. Krawetz and S. Misener (eds.). Bioinformatics Methods and Protocols: Methods in Molecular Biology. Humana Press, Totowa, N.J.
  20. Scott, K.D., P. Eggler, G. Seaton, M. Rosetto, E.M. Ablett, L.S. Lee, and R.J. Henry. 2000. Analysis of SSRs derived from grape ESTs. Theor. Appl. Genet. 100:723-726. https://doi.org/10.1007/s001220051344
  21. Semagn, K., A. Bjornstad, and M.N. Ndjiondjop. 2006. An overview of molecular marker methods for plants. African J. Biotechnol. 5:2540-2568.
  22. Simko, I. 2009. Development of EST-SSR markers for the study of population structure in lettuce (Lactuca sativa L.). J. Hered. 100:256-262. https://doi.org/10.1093/jhered/esn072
  23. Sneath, P.H.A. and R.R. Sokal. 1973. Numerical taxonomy: The principles and practice of numerical classification, Freeman W.H., San Francisco.
  24. Van de Wiel, C., P. Arens, and B. Vosman. 1999. Microsatellite retrieval in lettuce. Genome 42:139-149. https://doi.org/10.1139/g98-119
  25. Wang, F.G., H.L. Tian, J.R. Zhao, H.M. Yi, L. Wang, and W. Song. 2011. Development and characterization of a core set of SSR markers for fingerprinting analysis of Chinese maize varieties. Maydica 56:7-18.

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