Genetic Diversity and Species Identification of Cultivar Species in Subtribe Cucumerinae (Cucurbitaceae) Using RAPD and SCAR Markers

Juthaporn Saengprajak, Piyaporn Saensouk

Department of Biology, Faculty of Science, Mahasarakham University, Mahasarakham, Thailand..

DOI: 10.4236/ajps.2012.38131                

Abstract

DNA polymorphism in the cultivar species; Cucumis sativus L., C. melo L. and Benincasa hispida Cogn. of subtribe Cucumerinae (Cucurbitaceae) in the four northeastern provinces of Thailand was examined by using RAPD technique. Twenty 10-mer primers were produced 212 RAPD fragments, ranging from approximately 120 to 2531 bp. The genetic similarities were estimated from banding profiles using a NTSYS* version 2.1 as a basis for dendrogram construction via the UPGMA method. Cluster analysis divided the taxa under study into 2 clades. Moreover, a RAPD marker: Cm (OPJ11₇₀₀) was specified to C. melo, and this marker was converted into sequence characterized amplified region (SCAR) marker: Cm (SCJ11₅₁₆). A pair of sequence-specific primer of clones Cm (OPJ11₇₀₀) amplified a distinct single band of the same size as the RAPD clones. The SCAR marker was developed successfully to identify C. melo genotype.

Keywords

Cucumerinae; Cucurbitaceae; Genetic Diversity; Random Amplified Polymorphic DNA (RAPD); Sequence Characterized Amplified Region (SCAR); Species Identification

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1]	B. Sikdar, M. Bhattacharya, A. Mukherjee, A. Banerjee, E. Ghosh, B. Ghosh and S. C. Roy, “Genetic Diversity in Important Members of Cucurbitaceae Using Isozyme, RAPD and ISSR Markers,” Biologia Plantarum, Vol. 54, No. 1, 2010, pp. 135-140. doi:10.1007/s10535-010-0021-3
[2]	M. A. Helm and V. Hemleben, “Characterization of a New Prominent Satellite DNA of Cucumis metuliferus and Differential Distribution of Satellite DNA in Cultivated and Wild Species of Cucumis and in Related Genera of Cucurbitaceae,” Euphytica, Vol. 94, No. 2, 1997, pp. 219-226. doi:10.1023/A:1002976408213
[3]	A. Levi, C. E. Thomas, A. M. Simmons and J. A. Thies, “Analysis Based on RAPD and ISSR Markers Reveals Closer Similarities among Citrullus and Cucumis Species than with Praecitrullus fistulosus (Stocks) Pangalo,” Genetic Resources and Crop Evolution, Vol. 52, No. 4, 2005, pp. 465-472. doi:10.1007/s10722-005-2260-2
[4]	A. H. Paterson, S. D. Tanksley and M. E. Sorrells, “DNA Markers in Plant Improvement,” Advances in Agronomy, Vol. 46, 1991, pp. 39-90. doi:10.1016/S0065-2113(08)60578-7
[5]	M. Soller and J. S. Beckmann, “Genetic Polymorphism in Varietal Identification and Genetic Improvement,” Tag Theoretical Applied Genetics, Vol. 67, No. 1, 1983, pp. 25-33. doi:10.1007/BF00303917
[6]	K. Joshi, P. Chavan, D. Warude and B. Patwardhan, “Molecular Markers in Herbal Drug Technology,” Current Science, Vol. 87, No. 2, 2004, pp. 159-165.
[7]	J. K. W. Williams, A. R. Kubelik, K. J. Livak, J. A. Rafalski and S. V. Tingey, “DNA Polymorphisms Amplified by Arbitrary Primers Are Useful as Genetic Markers,” Nucleic Acids Research, Vol. 18, No. 22, 1990, pp. 6531-6535. doi:10.1093/nar/18.22.6531
[8]	M. A. Afzal, M. M. Haque and S. Shanmugasundaram, “Random Amplified Polymorphic DNA (RAPD) Analysis of Selected Mungbean (Vigna radiata L. Wilczek) Cultivars,” Asian Journal of Plant Sciences, Vol. 3, No. 1, 2004, pp. 20-24. doi:10.3923/ajps.2004.20.24
[9]	M. A. Karam, H. H. Elfayoumi and S. M. Mohamed, “Genetic Diversity and Taxonomic Status of Lotus glaber Mill Populations in Fayoum Depression,” Asian Journal of Plant Sciences, Vol. 5, No. 6, 2006, pp. 996-1001. doi:10.3923/ajps.2006.996.1001
[10]	A. Naz, Z. A. Swati and I. A. Khan, “Studies on Genetic Diversity in Pakistani Wheat Varieties Using Randomly Amplified Polymorphic DNA,” Pakistan Journal of Biological Sciences, Vol. 9, No. 15, 2006, pp. 2858-2861. doi:10.3923/pjbs.2006.2858.2861
[11]	P. W. Skroch, J. B. Dos Santos and J. Nienhuis, “Genetic Relationships among Phaseolus vulgaris Genotypes Based on RAPD Marker Data,” Annual Report of Bean Improvement Cooperation, Vol. 35, 1992, pp. 23-24.
[12]	B. K. Hoey, K. R. Crowe, V. M. Jones and N. O. Polans, “A Phylogenetic Analysis of Pisum Based on Morphological Characters, and Allozyme and RAPD Markers,” Tag Theoretical Applied Genetics, Vol. 92, No. 1, 1996, pp. 92-100. doi:10.1007/BF00222957
[13]	G. L. Brown-Guedira, J. A. Thompson, R. L. Nelson and M. L. Warburton, “Evaluation of Genetic Diversity of Soybean Introductions and North American Ancestors Using RAPD and SSR Markers,” Crop Science, Vol. 40, No. 3, 2000, pp. 815-823. doi:10.2135/cropsci2000.403815x
[14]	A. N. Kaga, N. Tomooka, Y. Egawa, K. Hosaka and O. Kamijima, “Species Relationships in the Subgenus Ceratotropis (Genus Vigna) as Revealed by RAPD Analysis,” Euphytica, Vol. 88, No. 1, 1996, pp. 17-24. doi:10.1007/BF00029261
[15]	I. Paran and R. W. Michelmore, “Development of Reliable PCR-Based Markers Linked to Downy Mildew Resistance Genes in Lettuce,” Tag Theoretical and Applied Genetics, Vol. 85, No. 8, 1993, pp. 985-993. doi:10.1007/BF00215038
[16]	S. L. Lee, J. Weber and R. Mohamed, “Genetic Variation and Molecular Authentication of Selected Aquilaria Species from Natural Populations in Malaysia Using RAPD and SCAR Markers,” Asian Journal of Plant Sciences, Vol. 10, No. 3, 2011, pp. 202-211. doi:10.3923/ajps.2011.202.211
[17]	A. Paul, S. Bandyopadhyay, P. Acharyya and S. S. Raychaudhuri, “Studies on Genetic Diversity of Twelve Accessions of Momordica charantia L. Using Morphological, RAPD and SCAR Markers,” Asian Journal of Plant Sciences, Vol. 9, No. 8, 2010, pp. 471-478. doi:10.3923/ajps.2010.471.478
[18]	J. Wang, Y.-H. Wai, F.-N. Ngan, P. P.-H. But and P.-C. Shaw, “Application of Sequence Characterized Amplified Region (SCAR) Analysis to Authenticate Panax Species and Their Adulterants,” Planta Medica, Vol. 67, No. 8, 2001, pp. 781-783. doi:10.1055/s-2001-18340
[19]	M. Y. Lee, E. J. Doh, H. P. Chae, H. K. Young, S. K. Eung, B. S. Ko and S.-E. Oh, “Development of SCAR Marker for Discrimination of Artemisia princeps and A. argyi from Other Artemisia Herbs,” Biological and Pharmaceutical Bulletin, Vol. 29, No. 4, 2006, pp. 629-663. doi:10.1248/bpb.29.629
[20]	A. Anuniwat, A. Chaveerach, T. Tanee and R. Sudmoon, “Development of SCAR Markers for Species Identification of the Genus Nepenthes (Nepenthaceae),” Pakistan Journal of Biological Sciences, Vol. 12, No. 22, 2004, pp. 1455-1461. doi:10.3923/pjbs.2009.1455.1461
[21]	M. Das, S. Bhattacharya and A. Pal, “Generation and Characterization of SCARs by Cloning and Sequencing of RAPD Products: A Strategy for Species-Specific Marker Development in Bamboo,” Annals of Botany, Vol. 95, No. 5, 2005, pp. 835-841. doi:10.1093/aob/mci088
[22]	J. J. Doyle and J. L. Doyle, “A Rapid DNA Isolation Procedure for Small Quantities of Fresh Leaf Tissue,” Phytochemical Bulletin, Vol. 19, 1987, pp. 11-15.
[23]	J. I. Hormaza, “Early Selection in Cherry Combining RAPDs with Embryo Culture,” Scientia Horticulturae, Vol. 79, No. 1-2, 1999, pp. 121-126. doi:10.1016/S0304-4238(98)00204-0
[24]	T. Mamiatis, E. F. Fritsch, J. Sambrook and J. Engel, “Molecular Cloning: A Laboratory Manual,” Cold Spring Harbor Laboratory Press, New York, 1982. doi:10.1002/abio.370050118
[25]	M. A. Serghini, C. Ritzenthaler and L. Pinck, “A Rapid and Efficient ‘Miniprep’ for Isolation of Plasmid DNA,” Nucleic Acids Research, Vol. 17, No. 9, 1989, p. 3604. doi:10.1093/nar/17.9.3604
[26]	J. Y. Um, H.-S. Chung, M.-S. Kim, H.-J. Na, H.-J. Kwon, J.-J. Kim, K.-M. Lee, S. J. Lee, J. P. Lim, K.-R. Do, W.-J. Hwang, Y.-S. Lyu, N.-H. An and H.-M. Kim, “Molecular Authentication of Panax ginseng Species by RAPD Analysis and PCR-RFLP,” Biological Pharmaceutical Bulletin, Vol. 24, No. 8, 2001, pp. 872-875. doi:10.1248/bpb.24.872
[27]	H.-T. Wolf, I. Zündorf, T. Winckler, R. Bauer and T. Dingermann, “Characterization of Echinacea Species and Detection of Possible Adulterations by RAPD Analysis,” Planta Medica, Vol. 65, No. 8, 1999, pp. 773-774. doi:10.1055/s-2006-960866
[28]	K. T. Chen, Y. C. Su, J. G. Lin, L. H. Hsin, Y. P. Su, C. H. Su, S. Y. Li, L. H. Cheng and S. J. Mao, “Identification of Atractylodes Plants in Chinese Herbs and Formulations by Random Amplified Polymorphic DNA,” Acta Pharmacologica Sinica, Vol. 22, No. 6, 2001, pp. 493-497.
[29]	S. Santhanalakshmi, S. Saikumar, S. Rao, A. SaiHarini, P. Khera, H. E. Shashidhar and P. Kadirvel, “Mapping Genetic Locus Linked to Brown Planthopper Resistance in Rice Oryza sativa L.,” International Journal of Plant Breeding and Genetics, Vol. 4, No. 1, 2010, pp. 13-22. doi:10.3923/ijpbg.2010.13.22
[30]	S. Bandyopadhyay and S. S. Raychaudhuri, “Development of ITS Based SCAR Markers for Some Medicinally Important Species of Phyllanthus,” Asian Journal of Plant Sciences, Vol. 9, No. 5, 2010, pp. 264-270. doi:10.3923/ajps.2010.264.270
[31]	A. H. Gorji, F. Darvish, M. Esmaeilzadehmoghadam and F. Azizi, “Application RAPD Technique for Recognition Genotypes Tolerant to Drought in Some of Bread Wheat,” Asian Journal of Biotechnology, Vol. 2, No. 3, 2010, pp. 159-168. doi:10.3923/ajbkr.2010.159.168
[32]	A. H. Amir, T. A. Malik , H. Raza and M. U. Rahman, “DNA Marker Studies for Leaf Nectaries in Upland Cotton,” Asian Journal of Plant Sciences, Vol. 1, No. 4, 2002, pp. 395-396. doi:10.3923/ajps.2002.395.396