Agrobacterium-mediated transformation of lettuce widely cultivated in Japan
2004
Nakano, M. (Niigata Univ. (Japan)) | Makino, M. | Sueyoshi, K.
ABSTRACT A simple and efficient system for producing transgenic plants via Agrobacterium-mediated transformation has been developed for crisp-head and loose-leaf type cultivars of lettuce (Lactuca sativa L.) widely cultivated in Japan. High efficiency of adventitious shoot regeneration was obtained from cotyledon explants on a medium containing 1 mg l**-1 NAA and 0.5 mg l**-1 BA. Cotyledons were co-cultivated for 3 days with A. tumefaciens strain EHA101/plG121Hm, which harbored the binary vector carrying the npt2, hpt and intron-containing gus (gus-intron) genes in the T-DNA region. Co-cultivated cotyledons were transferred onto a medium containing 1mg 1**-1 NAA, 0.5 mg 1**-1 BA, 50 mg 1**-1 kanamycin and 400 mg 1**-1 cefotaxime. Kanamycin-resistant adventitious shoots regenerated within 4 weeks were transferred to PGR-free medium containing 25 mg l-: kanamycin and 300 mg 1**-1 cefotaxime. Agrobacterium-free plantlets thus obtained were successfully transplanted to pots, and some of the plantlets showed GUS activity (GUS sup(+)) in their leaves. GUS sup(+) plantlets were verified to be transgenic by PCR analysis. Depending on the cultivar, 6 to 18 independent transgenic plants were obtained from 30 co-cultivated cotyledons.
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