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Comparison of DNA Extraction Protocols for Molecular Identification of Root Knot Nematode (Meloidogyne Spp.) Using Egg Masses

Authors:

W. M. S. U. K. Wijekoon,

University of Peradeniya, LK
About W. M. S. U. K.
Postgraduate Institute of Agriculture
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K. M. D. W. P. Nishantha,

Horticultural Crop Research and Development Institute, Gannoruwa, Peradeniya, LK
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D. M. De Costa

University of Peradeniya, LK
About D. M.
Postgraduate Institute of Agriculture
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Abstract

There is a dire need of alternative reliable and non-laborious methods with high resolution for identification of the root-knot nematodes at the species level where molecular techniques have shown the potentials. This study assessed four DNA extraction protocols, (i.e. TNES buffer method, Modified CTAB method, Phenol / Chloroform Method and 1% SDS method) to extract DNA from egg masses of root knot nematodes. Egg masses were collected from pure cultures of nematodes maintained in tomato cultivation under net house condition. After extraction of genomic DNA, Polymerase Chain Reaction was performed to determine the success of the method of DNA extraction. The 1% SDS method with incubation at -20 0C for 1hr and incubated at 65 0C for 1 hr and again incubated for 95 0C for 10 min, was a better method to result in intense PCR bands of the expected sizes (720 bp and 999 bp) when amplified by Meloidogyne genus and Meloidogyne incognita specific primers. This DNA extraction procedure could contribute as an effective method to molecular identification of species and other downstream applications of population studies of root knot nematodes.
How to Cite: Wijekoon, W.M.S.U.K., Nishantha, K.M.D.W.P. and De Costa, D.M., 2021. Comparison of DNA Extraction Protocols for Molecular Identification of Root Knot Nematode (Meloidogyne Spp.) Using Egg Masses. Tropical Agricultural Research, 32(3), pp.318–324. DOI: http://doi.org/10.4038/tar.v32i3.8495
Published on 30 Jul 2021.
Peer Reviewed

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