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Please use this identifier to cite or link to this item: http://hdl.handle.net/123456789/10951

Title: In-house protocol for isolation of restrictable and amplifiable genomic DNA from plants, fungi and bacteria.
Authors: Bahkali, A. H
Abd-Elsalam, K.A
Moslem, M.A
Al-Khedhairy, A. A
Keywords: 16S ribosomal RNA
biological materials
genomic DNA extraction
pathogenic bacteria
Issue Date: 2008
Publisher: Genes, Genomes and Genomics.
Citation: 2: 77-88
Abstract: Several rapid DNA isolation protocols are not available for plant and microorganisms with the same method. The method was applied to 5 plant species (tomato, cowpea, cotton, date palm and wild mint), 4 fungal species (Penicillium, Trichoderma, Fusarium and Rhizoctonia) and 4 bacterial species (Erwinia, Pseudomunas, Bacillius and Xanthomonas). Optimal extraction was achieved by incorporating an RNAse A and proteinase K enzymatic digestion step. The protocol produced an acceptable quantity of high-quality DNA. Up to 50 µg of DNA were routinely obtained from a minimal amount of 100 mg of fungal bacterial and plant sample. Fungal DNA prepared by this method was used as a template for PCR to amplify the internal transcribed spacer (ITS) and microsatellite-primed PCR and gave reproducible patterns. The amplicon length of the fragment ITS1/ITS4, ranged in size from 620 to 700 bp. The amplified PCR products of ITS regions in plants ranged in size from 550 to 700 bp, indicating polymorphisms of size in this region. The resultant amplicon was then incubated with the restriction endonucleases RsaI and CofI prior to analysis by gel electrophoresis. The protocol presented here offers an interesting and efficient alternative, eliminating most expensive kits, discounting the material cost per sample to $0.10.
URI: http://hdl.handle.net/123456789/10951
Appears in Collections:College of Science

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