A DNA Extraction Method Using a Silica-base Resin Type Kit for the Detection of Genetically Modified Papaya
Genetically modified (GM) papaya has not yet been approved for importation into, or cultivation in the European Union (EU) and Japan. A DNA extraction method using the Qiagen DNeasy Plant Mini Kit (PM method) and a method using a buffer containing cetyltrimethyl ammonium bromide (CTAB method) have b...
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| Published in | Food Hygiene and Safety Science (Shokuhin Eiseigaku Zasshi) Vol. 49; no. 2; pp. 63 - 69 |
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| Main Authors | , , , , , , , |
| Format | Journal Article |
| Language | English |
| Published |
Japan
Japanese Society for Food Hygiene and Safety
01.04.2008
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| Subjects | |
| Online Access | Get full text |
| ISSN | 0015-6426 1882-1006 |
| DOI | 10.3358/shokueishi.49.63 |
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| Abstract | Genetically modified (GM) papaya has not yet been approved for importation into, or cultivation in the European Union (EU) and Japan. A DNA extraction method using the Qiagen DNeasy Plant Mini Kit (PM method) and a method using a buffer containing cetyltrimethyl ammonium bromide (CTAB method) have been adopted as the official Japanese methods for detecting GM foods. However, the amounts of DNA extracted from papaya by these methods are very low. Therefore, we investigated an extraction method to obtain a high yield of DNA from raw or freeze-dried fresh papaya using the Promega Wizard DNA Clean-Up Resin System (WCR). The incubation for the extraction was carried out at 58°C without proteinase K for 15 min. The extract was applied to a mini-column, then the column was washed with 80% isopropyl alcohol, and genomic DNA adsorbed on the column was eluted with TE buffer. The WCR method gave a higher yield of genomic DNA, and was simpler and faster than the PM method or CTAB method. In addition, it could be used to extract genomic DNA from fresh papaya at various stages of ripeness. Based on these results, we propose that the present method using WCR is the most practical and useful way to extract genomic DNA for the purpose of detecting GM papaya. |
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| AbstractList | Genetically modified (GM) papaya has not yet been approved for importation into, or cultivation in the European Union (EU) and Japan. A DNA extraction method using the Qiagen DNeasy Plant Mini Kit (PM method) and a method using a buffer containing cetyltrimethyl ammonium bromide (CTAB method) have been adopted as the official Japanese methods for detecting GM foods. However, the amounts of DNA extracted from papaya by these methods are very low. Therefore, we investigated an extraction method to obtain a high yield of DNA from raw or freeze-dried fresh papaya using the Promega Wizard DNA Clean-Up Resin System (WCR). The incubation for the extraction was carried out at 58℃ without proteinase K for 15 min. The extract was applied to a mini-column, then the column was washed with 80% isopropyl alcohol, and genomic DNA adsorbed on the column was eluted with TE buffer. The WCR method gave a higher yield of genomic DNA, and was simpler and faster than the PM method or CTAB method. In addition, it could be used to extract genomic DNA from fresh papaya at various stages of ripeness. Based on these results, we propose that the present method using WCR is the most practical and useful way to extract genomic DNA for the purpose of detecting GM papaya. Genetically modified (GM) papaya has not yet been approved for importation into, or cultivation in the European Union (EU) and Japan. A DNA extraction method using the Qiagen DNeasy Plant Mini Kit (PM method) and a method using a buffer containing cetyltrimethyl ammonium bromide (CTAB method) have been adopted as the official Japanese methods for detecting GM foods. However, the amounts of DNA extracted from papaya by these methods are very low. Therefore, we investigated an extraction method to obtain a high yield of DNA from raw or freeze-dried fresh papaya using the Promega Wizard DNA Clean-Up Resin System (WCR). The incubation for the extraction was carried out at 58 degrees C without proteinase K for 15 min. The extract was applied to a mini-column, then the column was washed with 80% isopropyl alcohol, and genomic DNA adsorbed on the column was eluted with TE buffer. The WCR method gave a higher yield of genomic DNA, and was simpler and faster than the PM method or CTAB method. In addition, it could be used to extract genomic DNA from fresh papaya at various stages of ripeness. Based on these results, we propose that the present method using WCR is the most practical and useful way to extract genomic DNA for the purpose of detecting GM papaya. Genetically modified (GM) papaya has not yet been approved for importation into, or cultivation in the European Union (EU) and Japan. A DNA extraction method using the Qiagen DNeasy Plant Mini Kit (PM method) and a method using a buffer containing cetyltrimethyl ammonium bromide (CTAB method) have been adopted as the official Japanese methods for detecting GM foods. However, the amounts of DNA extracted from papaya by these methods are very low. Therefore, we investigated an extraction method to obtain a high yield of DNA from raw or freeze-dried fresh papaya using the Promega Wizard DNA Clean-Up Resin System (WCR). The incubation for the extraction was carried out at 58 degrees C without proteinase K for 15 min. The extract was applied to a mini-column, then the column was washed with 80% isopropyl alcohol, and genomic DNA adsorbed on the column was eluted with TE buffer. The WCR method gave a higher yield of genomic DNA, and was simpler and faster than the PM method or CTAB method. In addition, it could be used to extract genomic DNA from fresh papaya at various stages of ripeness. Based on these results, we propose that the present method using WCR is the most practical and useful way to extract genomic DNA for the purpose of detecting GM papaya.Genetically modified (GM) papaya has not yet been approved for importation into, or cultivation in the European Union (EU) and Japan. A DNA extraction method using the Qiagen DNeasy Plant Mini Kit (PM method) and a method using a buffer containing cetyltrimethyl ammonium bromide (CTAB method) have been adopted as the official Japanese methods for detecting GM foods. However, the amounts of DNA extracted from papaya by these methods are very low. Therefore, we investigated an extraction method to obtain a high yield of DNA from raw or freeze-dried fresh papaya using the Promega Wizard DNA Clean-Up Resin System (WCR). The incubation for the extraction was carried out at 58 degrees C without proteinase K for 15 min. The extract was applied to a mini-column, then the column was washed with 80% isopropyl alcohol, and genomic DNA adsorbed on the column was eluted with TE buffer. The WCR method gave a higher yield of genomic DNA, and was simpler and faster than the PM method or CTAB method. In addition, it could be used to extract genomic DNA from fresh papaya at various stages of ripeness. Based on these results, we propose that the present method using WCR is the most practical and useful way to extract genomic DNA for the purpose of detecting GM papaya. Genetically modified (GM) papaya has not yet been approved for importation into, or cultivation in the European Union (EU) and Japan. A DNA extraction method using the Qiagen DNeasy Plant Mini Kit (PM method) and a method using a buffer containing cetyltrimethyl ammonium bromide (CTAB method) have been adopted as the official Japanese methods for detecting GM foods. However, the amounts of DNA extracted from papaya by these methods are very low. Therefore, we investigated an extraction method to obtain a high yield of DNA from raw or freeze-dried fresh papaya using the Promega Wizard DNA Clean-Up Resin System (WCR). The incubation for the extraction was carried out at 58C without proteinase K for 15 min. The extract was applied to a mini-column, then the column was washed with 80% isopropyl alcohol, and genomic DNA adsorbed on the column was eluted with te buffer. The WCR method gave a higher yield of genomic DNA, and was simpler and faster than the PM method or CTAB method. In addition, it could be used to extract genomic DNA from fresh papaya at various stages of ripeness. Based on these results, we propose that the present method using WCR is the most practical and useful way to extract genomic DNA for the purpose of detecting GM papaya. Genetically modified (GM) papaya has not yet been approved for importation into, or cultivation in the European Union (EU) and Japan. A DNA extraction method using the Qiagen DNeasy Plant Mini Kit (PM method) and a method using a buffer containing cetyltrimethyl ammonium bromide (CTAB method) have been adopted as the official Japanese methods for detecting GM foods. However, the amounts of DNA extracted from papaya by these methods are very low. Therefore, we investigated an extraction method to obtain a high yield of DNA from raw or freeze-dried fresh papaya using the Promega Wizard DNA Clean-Up Resin System (WCR). The incubation for the extraction was carried out at 58°C without proteinase K for 15 min. The extract was applied to a mini-column, then the column was washed with 80% isopropyl alcohol, and genomic DNA adsorbed on the column was eluted with TE buffer. The WCR method gave a higher yield of genomic DNA, and was simpler and faster than the PM method or CTAB method. In addition, it could be used to extract genomic DNA from fresh papaya at various stages of ripeness. Based on these results, we propose that the present method using WCR is the most practical and useful way to extract genomic DNA for the purpose of detecting GM papaya. |
| Author | TSUCHIYA, Hisayo MAITANI, Tamio YAMADA, Toshiharu WATANABE, Takahiro SATOH, Shuji OHMORI, Kiyomi AKIYAMA, Hiroshi HIRAYAMA, Kuni |
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| References | 4) Peano, C., Samson, M. C., Palmieri, L., Gulli, M., Marmiroli, N. Qualitative and quantitative evaluation of the genomic DNA extracted from GMO and non-GMO foodstuffs with four different extraction methods. J. Agric. Food Chem., 52, 6962-6968 (2004). 1) Luthy, J., Brodmann, P., Buhler, H., Deu, J. Y., Eugster, A., Gafner, J. L., Hemmer, W., Hubner, P., Meyer, R., Pauli, U., Pfefferkorn, A., Rentsch, J., Ruf, J., Vogeli, U. Molekularbiologicsche Methoden. Swiss Food Manual, Chapter 52B (1998). 5) Wakui, C., Akiyama, H., Watanabe, T., Fitch, Maureen. M. M., Uchikawa., S., Yakahashi, K., Chiba, R., Fiji, A., Fujii, A., Hino, A., Maitani, T. A histochemical method using a substrate of β-glucronidase for detection of genetically modified papaya. J. Food Hyg. Soc. Japan, 45, 19-24 (2004). 2) Koopel, E., Stadler, M., Luthy, J., Hubner, P. Sensitive method for the detection of the genetically engineered soy bean <Roundup Ready>. Mitt. Gebiete Lebensm. Hyg., 88, 164-175 (1997). 3) Zimmermann, A., Luthy, J., Pauli, U. Quantitative and qualitative evalution of nine different extraction methods for nucleic acid on soya bean food samples. Z. Lebensm. Unters. Forsch. A, 207, 81-90 (1998). 1 2 3 4 WAKUI C (5) 2004; 45 |
| References_xml | – reference: 5) Wakui, C., Akiyama, H., Watanabe, T., Fitch, Maureen. M. M., Uchikawa., S., Yakahashi, K., Chiba, R., Fiji, A., Fujii, A., Hino, A., Maitani, T. A histochemical method using a substrate of β-glucronidase for detection of genetically modified papaya. J. Food Hyg. Soc. Japan, 45, 19-24 (2004). – reference: 1) Luthy, J., Brodmann, P., Buhler, H., Deu, J. Y., Eugster, A., Gafner, J. L., Hemmer, W., Hubner, P., Meyer, R., Pauli, U., Pfefferkorn, A., Rentsch, J., Ruf, J., Vogeli, U. Molekularbiologicsche Methoden. Swiss Food Manual, Chapter 52B (1998). – reference: 3) Zimmermann, A., Luthy, J., Pauli, U. Quantitative and qualitative evalution of nine different extraction methods for nucleic acid on soya bean food samples. Z. Lebensm. Unters. Forsch. A, 207, 81-90 (1998). – reference: 4) Peano, C., Samson, M. C., Palmieri, L., Gulli, M., Marmiroli, N. Qualitative and quantitative evaluation of the genomic DNA extracted from GMO and non-GMO foodstuffs with four different extraction methods. J. Agric. Food Chem., 52, 6962-6968 (2004). – reference: 2) Koopel, E., Stadler, M., Luthy, J., Hubner, P. Sensitive method for the detection of the genetically engineered soy bean <Roundup Ready>. Mitt. Gebiete Lebensm. Hyg., 88, 164-175 (1997). – ident: 2 – ident: 3 doi: 10.1007/s002170050299 – ident: 4 doi: 10.1021/jf040008i – ident: 1 – volume: 45 start-page: 19 issn: 0015-6426 issue: 1 year: 2004 ident: 5 doi: 10.3358/shokueishi.45.19 |
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| SubjectTerms | Carica - genetics Carica papaya DNA extraction DNA, Plant - isolation & purification Food Analysis - methods Food, Genetically Modified genetically modified (GM) papaya Genome, Plant - genetics Plants, Genetically Modified Polymerase Chain Reaction - methods Promega Wizard DNA Clean-Up Resin System qualitative PCR silica base resin type kit Silica Gel Silicon Dioxide |
| Title | A DNA Extraction Method Using a Silica-base Resin Type Kit for the Detection of Genetically Modified Papaya |
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