RAPD is a simple dominant marker system widely used in biology. Effectiveness of RAPD can be improved by selecting and redesigning primers whose priming sites occur in target sequence(s) of gene or organism at optimum distance. We developed software that uses sequences of random decamer primers and nucleotide sequence(s) as two input files. It locates the priming sites in input sequences and generates output files listing frequency and distance between priming sites. When the priming sites of a single primer occur more than once in a sequence with a distance of 200 to 2000?bp, the software also designs pairs of iSCAR primers. An input of 387 RAPD primers and 42,432 expressed sequences of oil palm are used as test. Wet-lab PCR results from a publication that used the same set of primers were compared with software output on priming sites. In the test sequences of oil palm covering 1.4% of genome, we found that at least 60% the primers chosen using software are sure of giving PCR amplification. We designed 641 iSCAR primers suitable for amplification of oil palm DNA. The software successfully predicted 92% (67 out of 73) of published polymorphic RAPD primers in oil palm. 1. Introduction The secret of differences between individual organism lies in their genetic material, called deoxyribonucleic acid (DNA). A genetic marker can be defined in one of the following ways: (a) a chromosomal landmark or allele that allows for the tracing of a specific region of DNA, (b) a specific piece of DNA with a known position on the genome, or (c) a gene whose phenotypic expression is usually easily discerned, used to identify an individual or a cell that carries it, or as a probe to mark a nucleus, chromosomes, or locus [1]. Genetic markers may not have a biological function, and they are inherited from one generation to next. Random amplified polymorphic (RAPD) DNA markers were introduced by Williams et al. [2] in 1990. RAPD markers can be implemented more rapidly and inexpensively than other type of markers. Prior knowledge of the DNA sequence for the targeted gene is not required, as the primers will bind somewhere in the sequence although exact location is unknown. RAPD primers are of decamer (10 base pairs) size and are randomly generated. The success of Polymerase Chain Reaction (PCR) is highly dependent on these short arbitrary oligonucleotides that hybridize onto the complementary DNA fragments. These short oligonucleotides function in pairs (one forward and one reverse primer) and are used to amplify [3] a set of DNA fragments. When choosing an arbitrary primer
References
[1]
R. C. King and W. D. Stansfield, A Dictionary of Genetics, Oxford University Press, New York, NY, USA, 1990, 4th edition.
[2]
J. G. K. 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, pp. 6531–6535, 1990.
[3]
M. Lexa, J. Horak, and B. Brzobohaty, “Virtual PCR,” Bioinformatics, vol. 17, no. 2, pp. 192–193, 2001.
[4]
K. Semagn, A. Bj?rnstad, and M. N. Ndjiondjop, “An overview of molecular marker methods for plants,” African Journal of Biotechnology, vol. 5, no. 25, pp. 2540–2568, 2006.
[5]
A. D. Ramessur and V. M. Ranghoo-Sanmukhiya, “RAPD marker-assisted identification of genetic diversity among mango (Mangifera indica) varieties in Mauritius,” International Journal of Agriculture and Biology, vol. 13, no. 2, pp. 167–173, 2011.
[6]
L. Bochra, Z. Nejia, L. Myriam, K. Karima, G. Abdelwahed, and M. Abdelaziz, “RAPD-based assessment of genetic diversity among annual caraway (Carum carvi) populations,” EurAsian Journal of BioSciences, vol. 5, pp. 37–47, 2011.
[7]
B. Y. Wang, L. Shi, Z. Y. Ruan, and J. Deng, “Genetic diversity and differentiation in Dalbergia sissoo (Fabaceae) as revealed by RAPD,” Genetics and Molecular Research, vol. 10, no. 1, pp. 114–120, 2011.
[8]
A. Saeed, I. Ahsan, N. Sehar, and A. Nisar, “Genetic diversity studies of coarse and fine rice using RAPD markers,” Frontiers of Agriculture in China, vol. 5, no. 2, pp. 129–134, 2011.
[9]
P. Vieira, W. Burgermeister, M. Mota, K. Metge, and G. Silva, “Lack of genetic variation of Bursaphelenchus xylophilus in Portugal revealed by RAPD-PCR analyses,” Journal of Nematology, vol. 39, no. 2, pp. 118–126, 2007.
[10]
S. Archana, D. Anish, M. Vindhya, and W. S. Lakra, “Molecular discrimination of six species of Bagrid catfishes from Indus river system using randomly amplified polymorphic DNA markers,” Molecular Biology Reports, vol. 38, no. 5, pp. 2961–2965, 2010.
[11]
C. Y. Fook, J. Y. Yap, T. L. Kiung, L. N. Z. Justina, W. K. Nelly, and V. K. Jayaraj, “Phylogenetic analysis of different breeds of domestic chickens in selected area of Peninsular Malaysia inferred from partial cytochrome b gene information and RAPD markers,” Animal Biotechnology, vol. 21, no. 4, pp. 226–240, 2010.
[12]
A. Milee, S. Neeta, and P. Harish, “Advances in molecular marker techniques and their applications in plant sciences,” Plant Cell Reports, vol. 27, no. 4, pp. 617–631, 2008.
[13]
K. F. Mahrous, H. A. I. Ramadan, S. H. Abdel-aziem, and M. A.-E. Mordy, “Genetic variations between camel breeds using microsatellite markers and RAPD techniques,” Journal of Applied Biosciences, vol. 39, pp. 2626–2634, 2011.
[14]
I. Paran and R. W. Michelmore, “Development of reliable PCR-based markers linked to downy mildew resistance genes in lettuce,” Theoretical and Applied Genetics, vol. 85, no. 8, pp. 985–993, 1993.
[15]
J. M. McDermott, U. Brandle, F. Dutly et al., “Genetic variation in powdery mildew of barley: development of RAPD, SCAR, and VNTR markers,” Phytopathology, vol. 84, no. 11, pp. 1316–1321, 1994.
[16]
J. Shi, J. Xin, and L. Xin, “A study on the RAPD and SCAR molecular markers of piper species,” Journal of Agriculture and Rural Development in the Tropics and Subtropics, vol. 110, no. 2, pp. 127–135, 2009.
[17]
A. W. Nurniwalis, N. Suhaimi, N. A. A. Siti, S. Aminah, and A. M. A. Mohamad, “Gene discovery via expressed sequence tags from the oil palm mesocarp,” Journal of Oil Palm Research, vol. 2, pp. 87–96, 2008.
[18]
C. L. Ho, Y. Y. Kwan, M. C. Choli et al., “Analysis and functional annotation of expressed sequence tags (ESTs) from multiple tissues of oil palm (Elaeis guineensis Jacq.),” BMC Genomics, vol. 8, p. 381, 2007.
[19]
J. J. Li, G. L. Pei, H. X. Pang, A. Bilderbeck, S. S. Chen, and S. H. Tao, “A new method for RAPD primers selection based on primer bias in nucleotide sequence data,” Journal of Biotechnology, vol. 126, no. 4, pp. 415–423, 2006.
[20]
D. J. Murphy, “Oil palm: future prospects for yield and quality improvements,” Lipid Technology, vol. 21, no. 11-12, pp. 257–260, 2009.
[21]
A. Beinaert and R. Vanderweyen, Contribution à l’étude Génétique et Biométrique des Variétés d’Elaeis Guineensis Jacq, Série Scientifique, l’institut National pour l’étude Agronomique du Congo Belge, 1941.
[22]
C. W. S. Hartley, The Oil Palm, Longman, London, UK, 2nd edition, 1988.
[23]
S. Bhasker and C. Mohankumar, “Association of lignifying enzymes in shell synthesis of oil palm (Elaeis guineensis Jacq.) fruit dura variety,” Indian Journal of Experimental Biology, vol. 39, no. 2, pp. 160–164, 2001.
[24]
A. Rival, L. Bertrand, T. Beulé, M. C. Combes, P. Trouslot, and P. Lashermes, “Suitability of RAPD analysis for the detection of somaclonal variants in oil palm (Elaeis guineensis Jacq),” Plant Breeding, vol. 117, no. 1, pp. 73–76, 1998.
[25]
D. Kambiranda, P. B. Subramani, and V. Padma, “Development of randomly amplified polymorphic DNA based SCAR marker for identification of Ipomoea mauritiana Jacq (Convolvulaceae),” Evidence-based Complementary and Alternative Medicine, vol. 2011, Article ID 868720, 6 pages, 2011.