Evaluation and optimization of DNA for RAPD analysis of Terminalia tomentosa

Mohd Rafi Wani¹ , Rajesh Rathore² , Javid Ahmad Malik³

Section:Research Paper, Product Type: Isroset-Journal
Vol.6 , Issue.1 , pp.275-278, Feb-2019

CrossRef-DOI:   https://doi.org/10.26438/ijsrbs/v6i1.275278

Online published on Feb 28, 2019

Copyright © Mohd Rafi Wani, Rajesh Rathore, Javid Ahmad Malik . This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
 

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Abstract :
Forest tree species are more demanding than other plants to extract a pure and high quality of DNA for molecular genetics. A modified protocol for Terminalia tomentosa genomic DNA extraction was developed by introducing modification on Cetyl Trimethyl Ammonium Bromide (CTAB) method. The principle modification we involved in the present study are; PVP (Polyvinylpyrrolidone), column based purification step and copper acetate solution. Fresh leaves were collected from the tender shoots of mature trees of Terminalia tomentosa. The yield and the purity of genomic DNA was monitored by measuring optical density (O.D.) at A260 and A280 with a Nanodrop Spectrophotometer (ND2000).The highest amount of genomic DNA with the best quality was isolated from column based step in the CTAB extraction buffer. The yield of DNA by this modified method was approximately 100-153 µg per 200mg of fresh leaf tissue. Both ratios were 2.1 and 1.8 respectively, indicating the absence of contaminating metabolites. The obtained DNA was then tested for suitability by random amplification of polymorphic DNA (RAPD)-PCR analysis. Clear band pattern were observed for DNA isolated from four accessions of Terminalia tomentosa, collected from different geographical locations of Achanakmar Amarkantak Biosphere Reserve, Central India. The results clearly indicate the utility of DNA for genetic diversity analysis of this forest tree species.

Key-Words / Index Term :
DNA extraction, CTAB method, quantity and purity, RAPD, Terminalia tomentosa

References :
[1] Sarwat M, Negi MS, Lakshmikumaran M, Tyagi AK, Das S, Srivastava PS. A standardized protocol for genomic DNA isolation from Terminalia arjuna for genetic diversity analysis. Electronic Journal of Biotechnology 9(1): 86-91,2006.
[2] Hanania U, Velcheva M, Sahar N, Perl A. An improved method for isolating high-quality DNA from Vitis vinifera nuclei. Plant Molecular Biology Reporter. 22(2): 173-177,2004.
[3] Crowley TM, Muralitharan MS, Stevenson TW. Isolating conifer DNA: a superior polysaccharide elimination method. Plant Molecular Biology Reporter. 21(1): 97a-97d,2003.
[4] Hills PN, Van SJ. An improved DNA extraction procedure for plant tissues with a high phenolic content. South African Journal of Botany. 68(4):549-550,2002.
[5] Shepherd M, Cross M, Stokoe RL, Scott LJ, Jones ME. High-throughput DNA extraction from forest trees. Plant Molecular Biology Reporter. 20(4): 425a-425j, 2002.
[6] Li YX, Su ZX, Chen F. Rapid extraction of genomic DNA from leaves and bracts of dove tree (Davidia involucrata). Plant Molecular Biology Reporter. 20(2): 185a-185e, 2002.
[7] Buldewo S, Jaufeerally-Fakim YF. Isolation of clean and PCR-amplifiable DNA from Anthurium andreanum. Plant Molecular Biology Reporter. 20(1): 71a-71g,2002.
[8] Pirttila AM, Hirsikorpi M, Kamarainen T, Jaakola L, Hohtola A. DNA isolation methods for medicinal and aromatic plants. Plant Molecular Biology Reporter. 19(3): 273a-273f,2001.
[9] Katterman FRH, Shattuck VI. An effective method of DNA isolation from the mature leaves of Gossypium species that contain large amounts of phenolics, terpeniodes and tannins. Preparative Biochemistry. 13: 347-359,1983.
[10] Bousquet J, Simon L, Lalonde M. DNA amplification from vegetative and sexual tissues of trees using polymerase chain reaction. Can. J. For. Res. 20: 254-257,1990.
[11] Pandey RN, Adams RP, Flournoy LE. Inhibition of random amplified polymorphic DNAs (RAPDs) by plant polysaccharides. Plant Mol. Biol. Rep., 14(1), 17–22,1996.
[12] Doyle JJ, Doyle JL. Isolation of plant DNA from fresh tissue. Focus, 12: 13–15, 1990.
[13] Krizman M, Baricevic D, Prosek M. Fast quantitative determination of volatile constituents in fennel by headspace-gas chromatography. Analytica Chimica Acta 557: 267-271,2006.
[14] Bokszczanin K, Prazybyla AA. Copper (II) acetate improves the quality of pear (Pyrus) DNA during extraction. Plant Mol. Biol. Rep., 24: 249a-249d,2006.
[15] Weishing K, Nybom H, Wolff K, Meyer W. DNA isolation and purification. In:DNA fingerprinting in plants and fungi, pp 44–59. CRC Press, Boca Raton, Florida,1995.
[16] Fang G, Hammer S, Grumet R. A quick and inexpensive method for removing polysaccharides from plant genomic DNA. Biotechniques, 13 (1): 52-54,1992.
[17] Sharma AD, Gill PK, Singh P. DNA isolation from dry and fresh samples of polysaccharide-rich plants. Plant Molecular Biology Reporter, 20(4): 415a-415f,2002.
[18] Loomis WD. Overcoming problems of phenolics and quinones in the isolation of plant enzymes and organelles. Methods in Enzymology, 31: 528-545,1974.
[19] Rogers SO, Benedich AJ. Extraction of DNA from milligram amounts of fresh, herbarium and mummified plant tissues. Plant Molecular Biology Report, 5(2): 69-76,1985.
[20] Dellaporta SL, Wood J, Hicks JB. A plant DNA miniprepration: version II. Plant Molecular Biology Report, 1: 19-21,1983.
[21] Khanuja SPS, Shasany AK, Darokar MP, Kumar S. Rapid isolation of DNA from dry and fresh samples of plants producing large amounts of secondary metabolites and essential oil. Plant Molecular Biology Reporter, 17(1): 74, 1999.
[22] Keb Llanes M, Gonzalez G, Chi-Manzanero B, Infante D. A rapid and simple method for small scale DNA extraction in Agavaceae and other tropical plants. Plant Molecular Biology Reporter, 20(3): 299a-299e, 2002.
[23] Sambrook J, Russel DW. Molecular cloning: a laboratory manual. Third Edition. Cold Spring Harbor Laboratory Press. New York,2001.