멜론 및 참외 순도 검정을 위한 SNP 마커 개발 및 F1 종자 순도 검정
SNP Marker Development for Purity Test of Oriental Melon and Melon
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멜론과 참외의 국내 소비 시장이 확대됨에 따라 다양한 F1품종이 개발되고 있다. 멜론과 참외의 F1 품종의 순도를 검정하기 위해 포장재배 등의 순도검정법이 이용되고 있으나 시간과 노력이 매우 많이 소요되기 때문에 분자마커를 이용한순도검정법의 개발이 필요하다. 본 연구에서는 멜론의 EST 염기정보로부터 30개의 SNP 프라이머 조합을 고안하여 멜론과 참외의 순도 검정을 위한 HRM분석방법을 개발하였다. 멜론 두 품종과 참외 한 품종의 양친 사이에 HRM 해리곡선의다형성을 보이는 10개의 마커를 선발하였으며 순도검정 마커를 선발하기 위해 blind test를 실시하였다. Blind test와 HRM 유전형 분석 결과가 일치하였으며 MEL SNP 2번과 12 마커를 이용하여 ‘레드 퀸’과 ‘얼쓰 VIP’의 F1 501개 개체에 대해순도검정을 실시하였다. HRM분석한 결과 모두 이형집합체로 나타나 100%의 순도를 보였다. 또한 HRM 방법을 이용하여 개발한 SNP 마커를 CAPS 마커로 전환하였다. CAPS 마커는 HRM 분석 마커와 비교하여 볼 때 멜론과 참외의 순도검정용 마커로 더 유용하게 활용될 수 있을 것으로 기대된다.
Abstract
Field screening method has been commonly used for purity test of F1 hybrid seeds in melon and oriental melon. However, as this method takes a lot of time and cost, molecular marker-based purity test is necessary. To develop molecular markers for purity test, thirty pairs of SNP (single nucleotide polymorphism) primers were obtained from melon EST sequences,and 10 polymorphic markers showing HRM (high resolution melting) polymorphisms between parents of two melon cultivars and one oriental melon cultivar were selected. Blind tests were performed to validate usefulness of the selected markers for purity test. Blind test results showed that HRM genotypes were matched with the expected identity of individual sample, F₁ hybrid, male or female parents. Three HRM-based SNP markers were converted to CAPS markers for general use which is favor to breeders. We expect that SNP markers developed in this study will be useful for purity test of F₁ hybrid seeds in melon and oriental melon.
참고문헌 (23)
- Ahmadian A, "Single-nucleotide polymorphism analysis by Pyrosequencing", Analytical Biochemistry, 280, pp.103~110 (2000)
- Ai CX, "Application of SSR markers in hybrid seed purity test of melon", Acta Horticulturae Sinica, 32(5), pp.902~904 (2005)
- Deleu W, "A set of EST-SNPs for map saturation and cultivar identification in melon", BMC Plant Biology, 9, pp.90~99 (2009)
- Gonzalez-Ibeas D, "MELOGEN: an EST database for melon functional genomics", BMC genomics, 8, pp.306~ (2007)
- Gundry CN, "Amplicon melting analysis with labeled primers: A closed-tube method for differentiating homozygotes and heterozygotes", Clin Chem, 49(3), pp.396~406 (2003)
- Howell WM, "Dynamic allele-specific hybridization: A new method for scoring single nucleotide polymorphisms", Nature Biotech, 17, pp.67~68 (1999)
- Kang WH, "Molecular mapping and characterization of a single dominant gene controlling CMV resistance in peppers (Capsicum annuum L.)", Theor Appl Genet, 120, pp.1587~1596 (2010)
- Li JF, "SSR markers for identification of purity of melon hybrids", Chinese J. of Agri Biotech, 5(3), pp.223~229 (2008)
- Liu JL, "Purity test of melon hybrid with SSR molecular markers", Mol Plant Breeding, 4(6(S)), pp.23~26 (2006)
- Ma GB, "Study on the application of starch gel electrophoresis for the identification of hybrid purity of watermelon and melon", Seed, 23(3), pp.30~32 (2004)
- McGuigan FE, "Single nucleotide polymorphism detection: allelic discrimination using TaqMan", Psychiatr Genet, 12, pp.133~136 (2002)
- Mhlanga M, "Using Molecular Beacons to detect single-nucleotide polymorphisms with real-time PCR", Methods, 25, pp.463~471 (2001)
- Montgomery J, "Simultaneous mutation scanning and genotyping by high resolution DNA melting analysis", Nature Protocols, 2, pp.59~66 (2007)
- Myakishev M, "High- Throughput SNP genotyping by allele-specific PCR with universal energy-transfer-labeled primers", Genome Res, 11, pp.163~169 (2001)
- Orita M, "Detection of polymorphisms of human DNA by gel electrophoresis as single-strand conformation polymorphisms", Proc Natl Acad Sci USA, 통권 : 86, pp.2766~2770 (1989)
- 박성우, "Optimization of High Resolution Melting Analysis and Discovery of Single Nucleotide Polymorphism in Capsicum", Horticulture, Environment, and Biotechnology, 50(1), pp.31~39 (2009), 한국원예학회
- Rapley R, "Molecular analysis and genome discovery", pp.113~126 (2004), Wiley
- Reed GH, "Sensitivity and specificity of single-nucleotide polymorphism scanning by high-resolution melting analysis", Clin Chem, 50, pp.1748~1754 (2004)
- Ming-Mao Sun, "SSR 마커에 의한 최근 육성 보급된 한국 벼 품종의 다양성과 품종 판별", 한국육종학회지, 41(2), pp.115~125 (2009), 한국육종학회
- Wang YH, "A genetic map of melon (Cucumis melo L.) based on amplified fragment length polymorphism (AFLP) markers", Theor Appl Genet, 95, pp.791~798 (1997)
- Wolf C, "PCR-RFLP analysis of mitochondrial DNA: a reliable method for species identification", J Agric Food Chem, 47, pp.1350~1355 (1999)
- Wu SB, "High resolution melting analysis of almond SNPs derived from ESTs", Theor Appl Genet, 118, pp.1~14 (2008)
- Zhai WQ, "Identification of purity of Hami melon hybrid with AFLP fingerprint", Acta Hort Sinica, 29(6), pp.587~ (2002)