Skip to main content
SpringerLink
Log in

Evaluation of High-Resolution Melting for Gene Mapping in Rice

  • Published:
Plant Molecular Biology Reporter Aims and scope Submit manuscript

Abstract

In this study, high-resolution melting (HRM) analysis was evaluated for gene mapping in rice with sequence-tagged site (STS) and simple sequence repeat (SSR) markers. A total of 103 out of 353 normal STS and SSR markers revealed polymorphic melting curves among the parental genotypes, and 12 of these were successfully used to genotype the F2 mapping population for HRM analysis. Additional electrophoresis findings demonstrated that HRM genotyping matched with traditional electrophoresis results. To optimize the HRM-marker screening efficiency, different HRM reaction conditions were evaluated. A 5-μl touchdown-polymerase chain reaction (PCR) system provided no significant improvement in screening efficiency but in a 10-μl touchdown-PCR system, the marker screening efficiency increased by 75%. Twenty-one markers were obtained for mapping purposes under the optimized reaction conditions. This study indicates that HRM analysis can speed up the gene mapping progress in rice, while saving a lot of manpower.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

References

  • Aguirre-Lamban J, Riveiro-Alvarez R, Garcia-Hoyos M, Cantalapiedra D, Avila-Fernandez A, Villaverde-Montero C, Trujillo-Tiebas MJ, Ramos C, Ayuso C (2010) Comparison of high-resolution melting analysis with denaturing high-performance liquid chromatography for mutation scanning in the ABCA4 gene. Invest Ophthalmol Vis Sci 51:2615–2619

    Article  PubMed  Google Scholar 

  • Akiyama H, Nakamura F, Yamada C, Nakamura K, Nakajima O, Kawakami H, Harikai N, Furui S, Kitta K, Teshima R (2009) A screening method for the detection of the 35 S promoter and the nopaline synthase terminator in genetically modified organisms in a real-time multiplex polymerase chain reaction using high-resolution melting-curve analysis. Biol Pharm Bull 32:1824–1829

    Article  PubMed  CAS  Google Scholar 

  • Araújo IS, De Souza Filho GA, Pereira MG, Faleiro FG, De Queiroz VT, Guimarães CT, Moreira MA, De Barros EG, Machado RCR, Pires JL, Schenell R, Lopes UV (2009) Mapping of quantitative trait loci for butter content and hardness in cocoa beans (Theobroma cacao L.). Plant Mol Biol Rep 27:177–183

    Article  Google Scholar 

  • Black WC 4th, Gorrochotegui-Escalante N, Duteau NM (2006) Heated oligonucleotide ligation assay (HOLA): an affordable single nucleotide polymorphism assay. J Med Entomol 43:238–247

    Article  PubMed  CAS  Google Scholar 

  • Chagné D, Gasic K, Crowhurst RN, Han YP, Bassett HC, Bowatte DR, Lawrence TJ, Rikkerink EHA, Gardiner SE, Korban SS (2008) Development of a set of SNP markers present in expressed genes of the apple. Genomics 92:353–358

    Article  PubMed  Google Scholar 

  • Chateigner-Boutin AL, Small I (2007) A rapid high-throughput method for the detection and quantification of RNA editing based on high-resolution melting of amplicons. Nucleic Acids Res 35:e114–e121

    Article  PubMed  Google Scholar 

  • Croxford AE, Rogers T, Caligari PD, Wilkinson MJ (2008) High-resolution melt analysis to identify and map sequence-tagged site anchor points onto linkage maps: a white lupin (Lupinus albus) map as an exemplar. New Phytol 180:594–607

    Article  PubMed  CAS  Google Scholar 

  • Erali M, Wittwer CT (2010) High resolution melting analysis for gene scanning. Methods 50:250–261

    Article  PubMed  CAS  Google Scholar 

  • Feng DS, Li Y, Wang HG, Li XF, Gao JR (2009) Isolation and evolution mode analysis of NBS–LRR resistance gene analogs from hexaploid wheat. Plant Mol Biol Rep 27:266–274

    Article  CAS  Google Scholar 

  • Galeano CH, Fernández AC, Gómez M, Blair MW (2009) Single strand conformation polymorphism based SNP and Indel markers for genetic mapping and synteny analysis of common bean (Phaseolus vulgaris L.). BMC Genomics 10:629–642

    Article  PubMed  Google Scholar 

  • Ganopoulos I, Argiriou A, Tsaftaris A (2010) Microsatellite high resolution melting (SSR-HRM) analysis for authenticity testing of protected designation of origin (PDO) sweet cherry products. Food Control 22:532–541

    Article  Google Scholar 

  • Gupta PK, Roy JK, Prasad M (2001) Single nucleotide polymorphisms: A new paradigm for molecular marker technology and DNA polymorphism detection with emphasis on their use in plants. Curr Sci 80:524–535

    CAS  Google Scholar 

  • Jaakola L, Suokas M, Häggman H (2010) Novel approaches based on DNA barcoding and high-resolution melting of amplicons for authenticity analyses of berry species. Food Chem 123:494–500

    Article  CAS  Google Scholar 

  • Korbie DJ, Mattick JS (2008) Touchdown PCR for increased specificity and sensitivity in PCR amplification. Nat Protoc 3:1452–1456

    Article  PubMed  CAS  Google Scholar 

  • Lehmensiek A, Sutherland MW, Mcnamara RB (2008) The use of high resolution melting (HRM) to map single nucleotide polymorphism markers linked to a covered smut resistance gene in barley. Theor Appl Genet 117:721–728

    Article  PubMed  CAS  Google Scholar 

  • Leng X, Xiao B, Wang S, Gui Y, Wang Y, Lu X, Xie J, Li Y, Fan L (2010) Identification of NBS-Type resistance gene homologs in tobacco genome. Plant Mol Biol Rep 28:152–161

    Article  CAS  Google Scholar 

  • Li B, Zhang DF, Jia GQ, Dai JR, Wang SC (2009) Genome-wide comparisons of gene expression for yield heterosis in maize. Plant Mol Biol Rep 27:162–176

    Article  CAS  Google Scholar 

  • Lu XJ, Jia YQ, Fan H, Zhang L, Jia J (2007) Methodological evaluation on PCR-DGGE technique in detecting DNA mutation and single nucleotide polymorphism. Sichuan Da Xue Xue Bao Yi Xue Ban 38:882–884

    PubMed  CAS  Google Scholar 

  • Mackay JF, Wright CD, Bonfiglioli RG (2008) A new approach to varietal identification in plants by microsatellite high resolution melting analysis: application to the verification of grapevine and olive cultivars. Plant Methods 4:8–17

    Article  PubMed  Google Scholar 

  • Mao F, Leung WY, Xin X (2007) Characterization of EvaGreen and the implication of its physicochemical properties for qPCR applications. BMC Biotechnol 7:76–91

    Article  PubMed  Google Scholar 

  • Margraf RL, Mao R, Highsmith WE, Holtegaard LM, Wittwer CT (2006) Mutation scanning of the RET protooncogene using high-resolution melting analysis. Clin Chem 52:138–141

    Article  PubMed  CAS  Google Scholar 

  • Prince JA, Feuk L, Howell WM, Jobs M, Emahazion T, Blennow K, Brookes AJ (2001) Robust and accurate single nucleotide polymorphism genotyping by dynamic allele-specific hybridization (DASH): design criteria and assay validation. Genome Res 11:152–162

    Article  PubMed  CAS  Google Scholar 

  • Reed GH, Wittwer CT (2004) Sensitivity and specificity of single-nucleotide polymorphism scanning by high-resolution melting analysis. Clin Chem 50:1748–1754

    Article  PubMed  CAS  Google Scholar 

  • Ronald PC, Albano B, Tabien R, Abenes L, Wu KS, Mccouch S, Tanksley SD (1992) Genetic and physical analysis of the rice bacterial blight disease resistance locus, Xa21. Mol Gen Genet 236:113–120

    PubMed  CAS  Google Scholar 

  • Tindall EA, Petersen DC, Woodbridge P, Schipany K, Hayes VM (2009) Assessing high-resolution melt curve analysis for accurate detection of gene variants in complex DNA fragments. Hum Mutat 30:876–883

    Article  PubMed  CAS  Google Scholar 

  • Tobler AR, Short S, Andersen MR, Paner TM, Briggs JC, Lambert SM, Wu PP, Wang YW, Spoonde AY, Koehler RT, Peyret N, Chen C, Broomer AJ, Ridzon DA, Zhou H, Hoo BS, Hayashibara KC, Leong LN, Ma CN, Rosenblum BB, Day JP, Ziegle JS, De La Vega FM, Rhodes MD, Hennessy KM, Wenz HM (2005) The SNPlex genotyping system: a flexible and scalable platform for SNP genotyping. J Biomol Tech 16:398–406

    PubMed  Google Scholar 

  • Ujino-Ihara T, Taguchi Y, Moriguchi Y, Tsumura Y (2010) An efficient method for developing SNP markers based on EST data combined with high resolution melting (HRM) analysis. BMC Res Notes 3:51–55

    Article  PubMed  Google Scholar 

  • Vossen RH, Aten E, Roos A, Den Dunnen JT (2009) High-resolution melting analysis (HRMA): more than just sequence variant screening. Hum Mutat 30:860–866

    Article  PubMed  CAS  Google Scholar 

  • Wu SB, Tavassolian I, Rabiei G, Hunt P, Wirthensohn M, Gibson JP, Ford CM, Sedgley M (2009) Mapping SNP-anchored genes using high-resolution melting analysis in almond. Mol Genet Genomics 282:273–281

    Article  PubMed  CAS  Google Scholar 

  • Wu SB, Wirthensohn MG, Hunt P, Gibson JP, Sedgley M (2008) High resolution melting analysis of almond SNPs derived from ESTs. Theor Appl Genet 118:1–14

    Article  PubMed  CAS  Google Scholar 

  • Yesilkaya H, Meacci F, Niemann S, Hillemann D, Rüsch-Gerdes S, LONG DRUG Study Group, Barer MR, Andrew PW, Oggioni MR (2006) Evaluation of molecular-Beacon, TaqMan, and fluorescence resonance energy transfer probes for detection of antibiotic resistance-conferring single nucleotide polymorphisms in mixed Mycobacterium tuberculosis DNA extracts. J Clin Microbiol 44:3826–3829

    Article  PubMed  CAS  Google Scholar 

  • Zhao F, Cai ZJ, Hu TZ, Yao HG, Wang L, Dong N, Wang B, Ru ZG, Zhai WX (2010) Genetic analysis and molecular mapping of a novel gene conferring resistance to rice stripe virus. Plant Mol Biol Rep 28:512–518

    Article  Google Scholar 

  • Zhao JT, Huang X, Chen YP, Chen YF, Huang XL (2009) Molecular cloning and characterization of an ortholog of NPR1 gene from Dongguan Dajiao (Musa spp. ABB). Plant Mol Biol Rep 27:243–249

    Article  CAS  Google Scholar 

  • Zhu WY, Lin J, Yang DW, Zhao L, Zhang YD, Zhu Z, Chen T, Wang CL (2009) Development of chromosome segment substitution lines derived from backcross between two sequenced rice cultivars, Indica recipient 93–11 and Japonica donor Nipponbare. Plant Mol Biol Rep 27:126–131

    Article  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by the Chinese New Genetically Modified Organism Varieties Programme (2009ZX08009-043B; 2009ZX08001-006B), the Chinese High-Tech Program (863 Plan, 2008AA02Z125), the Zhejiang Provincial Foundation for Natural Science (Z307451), and special grant from the Zhejiang Provincial Department of Science and Technology (2007 C12039). We are grateful to Professor Michael J. Adams, Rothamsted Research, Harpenden, UK for his correction of the English manuscript.

Author information

Authors and Affiliations

  1. College of Chemistry and Life Sciences, Zhejiang Normal University, Jinhua, 321004, People’s Republic of China

    Jinshan Li & Ruixian Dong

  2. State Key Laboratory Breeding Base for Zhejiang Sustainable Pest and Disease Control, MOA Key Laboratory for Plant Protection and Biotechnology, Zhejiang Provincial Key Laboratory of Plant Virology, Institute of Virology and Biotechnology, Zhejiang Academy of Agricultural Sciences, Hangzhou, 310021, People’s Republic of China

    Jinshan Li, Xuming Wang, Ruixian Dong, Yong Yang, Jie Zhou, Chulang Yu, Ye Cheng, Chengqi Yan & Jianping Chen

Authors
  1. Jinshan Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xuming Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ruixian Dong
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yong Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Jie Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Chulang Yu
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Ye Cheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Chengqi Yan
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Jianping Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jianping Chen.

Electronic Supplementary Material

Below is the link to the electronic supplementary material.

Supplemental Table 1

The 353 markers used for HRM analysis (XLS 91 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Li, J., Wang, X., Dong, R. et al. Evaluation of High-Resolution Melting for Gene Mapping in Rice. Plant Mol Biol Rep 29, 979–985 (2011). https://doi.org/10.1007/s11105-011-0289-2

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11105-011-0289-2

Keywords

Search

Navigation