The System Of In Vitro Culture And Shoot Regeneration From Leaves Of Pear Cultivars

The stable, efficient plant regeneration system is a key to success for transformation. In this study, the shoot tips isolated from Pyrus communis ' Bartlett' and ' Cornice', P. bretschneideri 'Zaosu' and P. pyrifolia 'Shenbuzhi' were used as explants. By means of the orthogonal experiment design, analysis of variance and multiple comparisons, the factors including genotypes, basical media, plant growth regulators, and supplements were analyzed and the system of in vitro culture and shoot regeneration from leaves of pear cultivars was established. The results were as follows:(1) The best sub-culture medium for in vitro perliferation of four cultivars was MS medium containing benzyladenine (BA) at 1. 0 mg/l and indolebutyric acid (IBA) at 0. lmg/l by examination of multiplication rate, differentiation shoots per microcutting, fully expanded leaves per microcutting. However, there were significant differences among pear varieties in shoot fresh and dry weight per explant.(2) From regenration rate, shoots per leaf and adventitious shoot regeneration efficiency, the optimal initiation medium for shoot regeneration of each cultivar was respectively as follows:For 'Bartlett', shoots regenerated best on MS supplemented with 0. 5 mg/l TDZ and 0. lmg/l IBA with regeneration rate of 62. 8%. For 'Cornice', 55. 6% leaves gave rise to shoots on the MS medium containing 3. 0 mg/l TDZ plus 0. lmg/l IBA. The highest regeneration frequency accounted for 75. 0% when cultured ' Zaosu' on the NN69 medium supplemented with 1.0 mg/l TDZ and 0. 4mg/l NAA. For 'Shenbuzhi', 62. 6~71. 4% regeneration rate were achieved from the NN69 medium supplemented with 1. 0~1.5 mg/l TDZ and 0. lmg/l NAA.55(3) On the best combination for leaves regeneration, AgNOs enhanced the regeneration efficiency of leaves while its concentration varied from 0.1mg/l to 0. 5 mg/l, from 0.1 to 2. 0 mg/litre, from 0. lmg/l to 0. 5 mg/l, from 0. lmg/l to 4. 0 mg/l, respectively for 'Bartlett','Cornice', 'Zaosu' and 'Shenbuzhi'. The most effective concentration of AgNO3 was 0. lmg/l for all cultivars but 'Bartlett'.(4) The shoot regeneration rate and efficiency were increasing along with the ratios of the nitrate: ammonium from 0:1 to 1:7 for ' Shenbuzhi' and 'Zaosu', and the best result was obtained from the 1:7 ratio of nitrate: ammonium. However, there was no adventitious bud appeared when the ratio of nitrate: ammonium was 0:1.(5 ) According to rooting rate, mean root number, mean root length, root fresh weight per shoot and root dry weight per shoot tested, the optimal rooting condition for four cultivars was respectively as follows:For ' Bartlett' , microcuttings were incubated the root iniation medium 1/4MS supplemented with IBA at 1.0mg/l and sucrose at 15.0g /l in light for 10 days, followed by 1/4MS containing IBA at 1. 0 mg/l, sucrose at 15.0g /l and 1.0g/l of AC.For 'Cornice', shoots were placed in the medium of 1/2MS supplemented with IBA at 5.0mg/l and sucrose at 15. 0 g /l in the light for 10 days, then into 1/2MS containing IBA at 5. 0 mg/l, sucrose at 15. 0 g /l and AC at 1. 0 g/l.For 'Zaosu', microcuttings were inoculated in MS containing IBA at 2. 5 mg/l and sucrose at 15.0g /lI under dark 10 days, and then transferred into root formation medium MS supplemented with sucrose at 15.0g/l and AC at 0. 5g/l without any growth regulators.For 'Shenbuzhi', shoots were first inoculated in 1/4MS containing IBA at 2. 5mg/l and sucrose at 5.0g /l in dark for 10 days, and 1/4MS supplemented with sucrose at 5.0g /l and AC at 0. 5g/l for the rest.