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 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.
Ohmori
Kiyomi
K
Chemistry Division, Kanagawa Prefectural Institute of Public Health, Chigasaki, Kanagawa, Japan.
Tsuchiya
Hisayo
H
Watanabe
Takahiro
T
Akiyama
Hiroshi
H
Maitani
Tamio
T
Yamada
Toshiharu
T
Hirayama
Kuni
K
Satoh
Shuji
S
D029441:Carica / Q000235:genetics; D018744:DNA, Plant / Q000302:isolation & purification*; D005504:Food Analysis / Q000379:methods*; D030841:Food, Genetically Modified; D018745:Genome, Plant / Q000235:genetics*; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction / Q000379:methods; D058428:Silica Gel; D012822:Silicon Dioxide
D018744:DNA, Plant; D058428:Silica Gel; D012822:Silicon Dioxide