In this study, I tried to establish the necessary protocols for the application of plant biotechnology to breeding of two Ipomoea species, I. batatas (sweet potato) and I. trichocarpa. The results obtained are summarized as follows. 1. High frequency of plant regeneration from leaf calli of sweet potato. High frequency plant regeneration was established from leaf calli of sweet potato (Ipomoea batatas (L.) Lam.) cv. Chugoku 25. Calli were formed from leaf segments on LS medium supplemented with 0.5mg/l 2,4-D, 3,000mg/l yeast extract, 5% (W/V) sucrose and 0.25% (W/V) gellan gum. Plant regeneration occurred at the frequency of more than 25% by transferring the calli onto the regeneration medium which was LS basal medium without any plant growth regulators. The presence of either abscisic acid (ABA) or silver nitrate (AgNO_3) in the callus induction medium promoted shoot regeneration. The optimum concentration was 2mg/l for both ABA and AgNO_3, the frequency of plant regeneration being 70 and 73.3%, respectively. 2. Plant regeneration from leaf calli of Ipomoea trichocarpa Ell. Calli were induced from leaf nieces of Ipomoea trichocarpa on the LS basal medium supplemented with 0.5mg/l 2,4-D, 5mg/l ABA, 3g/l yeast extract, 5% sucrose and 0.2% gellan gum. These calli formed adventitious buds on the regeneration media containing more than 2mg/l BA. The highest adventitious bud formation was obtained at 10mg/l BA. Adventitious roots were also produced from calli on all kinds of the regeneration media used in the present study. There are two type of the adventitious roots from leaf calli, thick roots (1.0mm in diameter) and thin roots (less than 0.2mm in diameter). When the calli with adventitious roots were transferred to plant growth regulator (PGR)-free LS medium, 63.3 - 90.6% of the roots produced shoots. Shoots were only regenerated from the thick roots. It was suggested that some adventitious shoot primordia were already developed on the thick roots. Shoots regenerated from both leaf calli and adventitious roots could subsequently grow to plantlets. Then these plantlets were transplanted to pots containing a mixture of vermiculite and perlite. All of them grew normally in the growth chamber and had normal seed fertility. No morphological differences were observed in 71 regenerated plants. 3. Callus and root formation from protoplasts of mesophyll and cultured cells. Protoplasts were isolated from mesophyll tissue, callus and suspension cell cultures of sweet potato. Viable protoplasts (7.1×10^4-8.6×10^5 protoplasts/g fresh weight) were obtained from the third to fifth leaves from the top of these plants, whereas younger ones, the first and the second, yielded only small and fragile protoplasts which dld not divide in the medium. When chopped leaf tissue was incubated in sterile distilled water for 16 h prior to enzymatic digestion, the vield of protoplasts showed 20-fold increased compared with untreated material in all of the cultivars tested in the present study. In protoplast isolation from cultured cell, freshly isolated protoplasts from cultured cells were cytoplasmic-rich and 30 to 40μm in diameter. The yield of protoplast was 1-2×10^6 cells/g fresh wight in both callus and cell suspension cultures. No clear difference in the yield of protoplasts was observed between two cultivars. About 13% of mesophyll protoplasts and 25% of cultured cell protoplasts divided on the third day. When these calli derived from both mesophyll and callus cells protoplasts were transferred onto the regeneration media, a few roots were formed from the calli placed on the media containing both BA at 0.5, 1.0 or 2.0 mg/l and IAA at 0.2 mg/l, but no shoots were produced. 4. Transformation of sweet potato plants by Agrobacterium rhizogenes. Transgenic sweet potato plants were obtained after Agrobacterium rhizogenes-mediated transformation. Leaf disks of in vitro plants were inoculated with different A. rhizogenes strains. Numerous hairy roots were
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