PRODUCTION OF HIGH-DIGESTIBLE,BROWN MIDRIB TYPE FORAGE

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In order to produce the high-digestible, as it were, the brown midrib type forage contained less lignin, we carried out the experiments as follows ;1) Bird's-foot trefoil (Lotus corniculatus L.) are transformed with Agrobacterium tumefaciens carrying the plasmid vector pBI121 contained a kanamycin resistant gene (NTP II) and beta-glucuronidase gene (GUS). Resistant plants were regenerated at high frequency from green calli developed on the inoculated hypocotyl cuttings under the kanamycin selection.2) The histological GUS assay showed that 7.0% of the resistant shoots also expressed the GUS gene in a variety tissues. The stable integration of this gene was confirmed by polymerase chain reaction (PCR) analysis.3) A partial gene encoding CAD (Cinnamyl Alcohol Dehydrogenase) was isolated from Aralia cordata using PCR.An 1.2-kb portion of the sequence of the CAD gene obtained by PCR was inserted into an antisense orientation between the 35S promoter and the NOS terminater gene in T-DNA region of pGAH/SL-GUS.This constructed plasmid pGAH/anti-UCAD was introduced into bird's foot trefoil. The transgenic plants were selected by means of the resistance to kanamycin and hygromycin phosphotransferase.4) The stable integration of the antisense gene was confirmed by PCR in the transgenic plants. The CAD activity of the transgenic plants was lower significantly than that of the control plants, but the lignin and the aldehyde contents and the in vitro digestibility by rumen mincroorganisms were not significantly different between the transgenic and control plants.