Study On Technique Tissue Culture In Phellodendron Chinense Schneid

In this paper, a plant regeneration system for Phellodendron chinense Schneid was developed by using various explants including immature embryos, shoot tips, young leaves and leafstalks, and shoots, leaves and roots regenerated from immature embryo in vitro culture. The best culture medium for different explants, the kinds and concentrations of plant growth regulators, the influence of H3BO3 and AC on rooting were investigated by means of orthogonal experiment design, full combination experiment design, and one-factor experiment design and analysis of variance and multiple comparisons. The results were as follows:1. The proper embryo germination medium was 1/2MS+6-BA0.2mg/L+KT0.2 mg/L +NAA0.02 mg/L +IBA0.02 mg/L in which the rate of germination was 72.50%.2. The best medium for initial shoot multiplication was MS+6-BA1.0 mg/L +NAA0.1 mg/L, and foe later multiplication, the best multiplication medium was MS+6-BA0.2 mg/L +NAA0.02 mg/L, in which the multiplication rates were 4.50 and 4.16 and the shoot heights were 3.4cm and 3.6cm respectively.3. The best medium for inducing callus from roots obtained from embryo culture was MS+6-BA0.5 mg/L +KT0.2 mg/L +IBA0.1 mg/L +NAA0.5 mg/L, in which the rate of callus formation was 100%. The best medium for redifferentiation of callus was MS+6-BA1.0 mg/L +NAA0.1 mg/L+GA₃1.0, in which the rate of redifferentiation was 25.00% with the average shoot number being 3.36 per explant and the average shoot height being 1.8cm.4. The best medium for inducing callus from leaves which were from embryo culture was 1/2MS+6-BA1.0 mg/L +2,4-D0.1 mg/L, where the formed calluses were green and compact, and the rate of callus formation was 78.56%. The best medium for redifferentiation of callus was MS+6-BA0.2 mg/L +NAA0.02 mg/L, in which the rate of redifferentiation was 37.78% with 3⁵ shoots being obtained on each callus explant. It took about 60 days to form 2cm-long shoots.5. The best medium for initial culture of shoot-tips was MS+6-BA0.5 mg/L +NAA0.05 mg/L, in which the formation rate of well developed shoots was 77.78%, and the rate of vitrification was very low. The best medium for multiplication was MS+6-BA0.5 mg/L+NAA0.05 mg/L +GA3I.O mg/L, in which the axillary buds sprouted well.6. The best medium for inducing callus from leaves on young shoot was MS+6-BA0.5 mg/L +NAA0.3 mg/L, on which the rate of forming green and compact callus was 76.97%, and there were no vitrification and browning. The best medium for redifferentiation of callus was MS+6-BA2.0 mg/L +NAA0.1 mg/L, on which the rate of redifferentiation was 63.89%. A large number of bud spots were formed on the callus and after 60 days' culture they developed to shoots of about 2.0cm in length, but the shoots were slim and a little vitreous.7. The best medium for inducing callus from leafstalks on young shoots was MS+6-BA0.3 mg/L +NAA0.7 mg/L, the rate of forming green and compact callus was 89.44%. The calluses were first cultured on MS+6-BA0.8 mg/L +NAA0.05 mg/L for 20 days and then on MS+6-BA2.0 mg/L +NAA0.3 mg/L for 20 days, resulting in about 20 shoots being regenerated on each callus explant and the shoot height being about 3cm. The best medium for shoot multiplication was MS+6-BA0.6 mg/L +NAA0.09 mg/L, on which the rate of multiplication was 6.47.8. The best medium for the rooting of the shoots in immature embryo culture was 1/2MS+NAA0.05 mg/L +IBA0.05 mg/L +AC0.3%+H3BO314.0 mg/L, on which the rate of rooting was 85.42%. The best medium for the rooting of the shoots obtained from other explants was 1/2MS+NAA0.1 mg/L +AC0.3%+ H3BO314.0 mg/L, in which the rate of rooting was 62.50%.9. The best transplanting medium was the moisted compost of humus and sand, in which 94.44% were successfully transplanted.