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High resolution melting analysis as a rapid and highly sensitive method for Cichorium plasmotype characterization

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Abstract
Somatic hybridization in Cichorium species has already been extensively investigated. Hybrid or cybrid characterization requires an effective plasmotype screening method. We evaluated high resolution melting (HRM) analysis for the detection of specific mitochondrial (mt) and chloroplast (cp) markers to distinguish two industrial chicory (Cichorium intybus var. sativum) plasmotypes from five wild type chicory (C. intybus) and two endive (C. endivia) plasmotypes. Three mt (coxII-2, cob-1, and cob-2) and three cp HRM markers (trnL-trnF, trnL-trnF-2, and ndhF-1) were successfully developed. Two markers (coxII-2 and trnL-trnF) were additionally able to discriminate heterozygous plasmotypes containing at least 25 % of one parental plasmotype. Moreover, the technique was successfully used to characterize the cytoplasms of 50 simple sequence repeats (SSR) confirmed somatic hybrids of C. intybus var. sativum ‘VL52’ and C. endivia var. crispum ‘Wallone Despa’. HRM enables a rapid (less than 2 h) and efficient high-throughput scanning of multiple fragments due to the 384-well plates and is cost efficient because of its small reaction volume of 10 μl. This is the first report on the use of HRM analysis on Cichorium species. The technique is a fast and simple alternative for laborious and costly sequencing in plasmotyping regenerants obtained after somatic fusion.
Keywords
Cytoplasmic markers, High resolution melting (HRM), Somatic hybridization, Cichorium, SOMATIC HYBRID, INTYBUS L, IDENTIFICATION, MARKERS, SNP, DNA, POLYMORPHISMS, REGENERATION, AMPLICONS, CHICORY

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MLA
Deryckere, Dieter, Ellen De Keyser, Tom Eeckhaut, et al. “High Resolution Melting Analysis as a Rapid and Highly Sensitive Method for Cichorium Plasmotype Characterization.” PLANT MOLECULAR BIOLOGY REPORTER 31.3 (2013): 731–740. Print.
APA
Deryckere, D., De Keyser, E., Eeckhaut, T., Van Huylenbroeck, J., & Van Bockstaele, E. (2013). High resolution melting analysis as a rapid and highly sensitive method for Cichorium plasmotype characterization. PLANT MOLECULAR BIOLOGY REPORTER, 31(3), 731–740.
Chicago author-date
Deryckere, Dieter, Ellen De Keyser, Tom Eeckhaut, Johan Van Huylenbroeck, and Erik Van Bockstaele. 2013. “High Resolution Melting Analysis as a Rapid and Highly Sensitive Method for Cichorium Plasmotype Characterization.” Plant Molecular Biology Reporter 31 (3): 731–740.
Chicago author-date (all authors)
Deryckere, Dieter, Ellen De Keyser, Tom Eeckhaut, Johan Van Huylenbroeck, and Erik Van Bockstaele. 2013. “High Resolution Melting Analysis as a Rapid and Highly Sensitive Method for Cichorium Plasmotype Characterization.” Plant Molecular Biology Reporter 31 (3): 731–740.
Vancouver
1.
Deryckere D, De Keyser E, Eeckhaut T, Van Huylenbroeck J, Van Bockstaele E. High resolution melting analysis as a rapid and highly sensitive method for Cichorium plasmotype characterization. PLANT MOLECULAR BIOLOGY REPORTER. 2013;31(3):731–40.
IEEE
[1]
D. Deryckere, E. De Keyser, T. Eeckhaut, J. Van Huylenbroeck, and E. Van Bockstaele, “High resolution melting analysis as a rapid and highly sensitive method for Cichorium plasmotype characterization,” PLANT MOLECULAR BIOLOGY REPORTER, vol. 31, no. 3, pp. 731–740, 2013.
@article{3205940,
  abstract     = {Somatic hybridization in Cichorium species has already been extensively investigated. Hybrid or cybrid characterization requires an effective plasmotype screening method. We evaluated high resolution melting (HRM) analysis for the detection of specific mitochondrial (mt) and chloroplast (cp) markers to distinguish two industrial chicory (Cichorium intybus var. sativum) plasmotypes from five wild type chicory (C. intybus) and two endive (C. endivia) plasmotypes. Three mt (coxII-2, cob-1, and cob-2) and three cp HRM markers (trnL-trnF, trnL-trnF-2, and ndhF-1) were successfully developed. Two markers (coxII-2 and trnL-trnF) were additionally able to discriminate heterozygous plasmotypes containing at least 25 % of one parental plasmotype. Moreover, the technique was successfully used to characterize the cytoplasms of 50 simple sequence repeats (SSR) confirmed somatic hybrids of C. intybus var. sativum ‘VL52’ and C. endivia var. crispum ‘Wallone Despa’. HRM enables a rapid (less than 2 h) and efficient high-throughput scanning of multiple fragments due to the 384-well plates and is cost efficient because of its small reaction volume of 10 μl. This is the first report on the use of HRM analysis on Cichorium species. The technique is a fast and simple alternative for laborious and costly sequencing in plasmotyping regenerants obtained after somatic fusion.},
  author       = {Deryckere, Dieter and De Keyser, Ellen and Eeckhaut, Tom and Van Huylenbroeck, Johan and Van Bockstaele, Erik},
  issn         = {0735-9640},
  journal      = {PLANT MOLECULAR BIOLOGY REPORTER},
  keywords     = {Cytoplasmic markers,High resolution melting (HRM),Somatic hybridization,Cichorium,SOMATIC HYBRID,INTYBUS L,IDENTIFICATION,MARKERS,SNP,DNA,POLYMORPHISMS,REGENERATION,AMPLICONS,CHICORY},
  language     = {eng},
  number       = {3},
  pages        = {731--740},
  title        = {High resolution melting analysis as a rapid and highly sensitive method for Cichorium plasmotype characterization},
  url          = {http://dx.doi.org/10.1007/s11105-012-0547-y},
  volume       = {31},
  year         = {2013},
}

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