Abstract: [Objective]Apple rootstocks are used to influence precocity, tree size, fruit quality, yield efficiency, mineral uptake, apple replant disease, and to withstand adverse environmental conditions. Especially dwarfing rootstocks play an important role in developing modern orchards. New apple rootstock cultivar ‘Luzhen 1’ was a semi-dwarf rootstock which was selected and released by Shandong Institute of Pomology. This new rootstock produces trees similar in size to M.26 and it can be propagated clonally, and it has good graft union compatibility with many scion cultivars. The purpose of this paper is to establish technological system of high-efficiency shoot regeneration from leaf explants of new cultivar ‘Luzhen 1’, to provide basis for commercial propagation of plants and further genetic improvement using biotechnological methods. [Methods]In miD-to-late April, semi-lignified healthy shoots of ‘Luzhen 1’ were used to culture in vitro sterile plantlets. The leaves from secondary sterile plants were induced to adventitious shoots. Through the above experiments, the effects of carbon source, type and concentration of cytokinin on leaf regeneration were studied. Further, the effects of basic medium and sucrose concentration on rooting of adventitious shoots were also studied. [Results]The axillary buds on semi-lignified shoots germinated and extended to form sterile green plantlets, after culturing on the axillary bud initiation medium for 3 weeks. These plantlets grew well on the secondary proliferation medium, showing both good proliferation and good elongation growth, and the monthly multiplication ratio was above 5.0, indicating the easy multiplication and propagation of ‘Luzhen 1’. The two carbon sources tested, D-sorbitol and sucrose, were both effective in inducing adventitious bud regeneration. Except that the adventitious bud regeneration rate on D-sorbitol was significantly higher than that on sucrose at a lower concentration of 0.6 mg·L-1 of cytokinin TDZ, there was no significant difference in the adventitious bud regeneration rate between the two carbon sources in other treatments, both of which were as high as 90%. For the average number of adventitious buds per leaf, there was no significant difference between the two carbon sources under the same type and concentration of cytokinin. These results indicate that the carbon source is not strictly required for adventitious bud regeneration of isolated leaves of the new stock. Under the same carbon source condition, the adventitious bud regeneration rate and average bud number per leaf were the lowest when TDZ was at a lower concentration of 0.6 mg·L-1, and the results were consistent for the two carbon sources. When the carbon source in the medium was D-sorbitol, there was no significant difference in the regeneration rate of adventitious buds between TDZ and BA, and between different concentrations. However, the average number of adventitious buds per leaf at 1 mg·L-1 concentration of TDZ was significantly higher than that at 0.6 mg·L-1, while there was no significant difference between the two concentrations of BA. When the carbon source in the medium was sucrose, the regeneration rate of TDZ at a lower concentration of 0.6 mg·L-1 was significantly lower than that of other treatments, but the average number of adventitious buds per leaf was not significantly different among different treatments. On adventitious-bud induction medium, cytokinin BA was more easily than TDZ to directly induce the formation of elongated and growing adventitious shoots, while adventitious buds induced by TDZ did not easily obtain these shoots on the induction medium, and needed to be transferred to the proliferation medium without TDZ. The best medium combination for inducing adventitious bud regeneration in leaves is MS medium with 1 mg·L-1 TDZ, 0.3 mg·L-1 IBA and 30 g·L-1 sucrose. After rooting induction culture for 12 days, the emergence of early adventitious root protrusions could be observed. After culture for 18 days, the formation of obvious short roots, more than 88% of the final roots, could be observed. After culture for 22 days, no new adventitious roots could be generated. After that, with the extension of culture time, the change was mainly the root elongation growth. All the eight root medium tested could successfully induce adventitious shoot rooting, but the rooting rate varied greatly, and the average number of roots per plant had no significant difference. Independent of the concentration of IBA, the basic medium 1/4 MS with lower sucrose (20 g·L-1) was more conducive to improving the rooting rate and the average number of roots per plant, than with higher sucrose (30 g·L-1). In addition to 0.3 mg·L-1 IBA, rooting rate on 20 g·L-1 sucrose was significantly higher than that on 30 g·L-1 sucrose. On the basic medium 1/2 MS, there was no significant difference in rooting rate and number of roots per plant, no matter between two sucrose concentrations or two IBA concentrations. The rooting rate of the four treatments could reach more than 70%, indicating that on the basic medium 1/2 MS, the requirement of sucrose concentration and IBA concentration for the adventitious rooting of 'Luzhen 1' was not strict. These indicated that adventitious shoots of ‘Luzhen 1’ were easy to root. The optimal rooting medium was 1/4 MS supplemented with 0.3~0.5 mg·L-1 IBA and 20 g·L-1 sucrose. The rooting rate was over 93% and the average number of roots per plant was 5.8~6.0. [Conclusions]It was relatively easy to induce bud regeneration from leaf explants and in vitro rooting from adventitious shoots of apple semi-dwarf rootstock new cultivar ‘Luzhen 1’. The best adventitious bud induction medium of ‘Luzhen 1’ was MS+1 mg·L-1 TDZ+0.3 mg·L-1 IBA+30 g·L-1 sucrose. The best medium of in vitro rooting medium from adventitious shoots of ‘Luzhen 1’ was 1/4MS+0.3~0.5 mg·L-1IBA+20 g·L-1sucrose.
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