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Title: A HISTOCHEMICAL METHOD USING A SUBSTRATE OF BETA-GLUCURONIDASE FOR DETECTION OF GENETICALLY MODIFIED PAPAYA

Author
item WAKUI, C.
item AKIYAMA, H.
item WATANABE, T.
item Fitch, Maureen
item UCHIKAWA, S.
item KI, M.
item TAKAHASHI, K.
item CHIBA, R.
item FUJII, .
item HINO, A.
item MAITANI, T.

Submitted to: Trade Journal Publication
Publication Type: Trade Journal
Publication Acceptance Date: 10/15/2003
Publication Date: 2/1/2004
Citation: Wakui, C., Akiyama, H., Watanabe, T., Fitch, M.M., Uchikawa, S., Ki, M., Takahashi, K., Chiba, R., Fujii, .., Hino, A., Maitani, T. 2004. A histochemical method using a substrate of beta-glucuronidase for detection of genetically modified papaya. Journal of Food Hygenics Society of Japan. 45 (1): 19-24.

Interpretive Summary: Abstract only.

Technical Abstract: A histochemical assay for detecting genetically modified (GM) papaya (derived from Line 55-1) is described. GM papayas, currently undergoing a safety assessment in Japan, was developed using a construct that included a ß-glucuronidase (GUS) reporter gene linked to a virus coat protein (CP) gene. Histochemical assay was used a visualize the blue GUS reaction product from transgenic seed embryos. Twelve embryos per fruit were extracted from the papaya seeds using a surgical knife. The embryos were incubated with the substrate 5-bromo-4-chloro-3-indolyl- ß-D-glucuronide (X-Gluc) in a 96-well microtiter plate for 10-15 hours at 37oC. Seventy-five percent of GM papaya embryos should turn blue theoretically. The histochemical assay results were completely consistent with those from a qualitative polymerase chain reaction (PCR) method developed by this laboratory. Furthermore, the method was validated in a five-laboratory study. The method for detection of GM papaya is rapid and simple, and does not require use of specialized equipment.