Enhancing Micropropagation Efficiency of Apple Rootstocks by Identifying Limitations of Microshoot Elongation

During the initial phase of shoot culture, in vitro shoot growth of apple rootstocks is stunted which is probably related to bud dormancy. To overcome the limitations of stunted in vitro shoot growth in micropropagation, the effects of plant growth regulators, chilling nodal cuttings, and light quality on in vitro shoot growth were investigated. Gibberellin (GA3) did not significantly improve shoot elongation, but 6- benzylaminopurine (6-BA) improved shoot proliferation and elongation compared with thidiazuron (TDZ), zeatin (ZT) or indole-3-butyricacid (IBA). Chilling for at least 4 weeks improved shoot growth; six weeks chilling was optimum for G.30 rootstock shoot growth. However, chilling for 8 weeks suppressed shoot growth compared with 6 weeks. Shoot explants collected in April and chilled for 0, 4, or 6 weeks produced more and longer shoots compared with August or November, excluding chilling for 8 weeks which had similar shoot growth. Red light significantly improved shoot growth, and the effect was enhanced when combined with 0.5 mg/L GA3. Blue light decreased the shoot growth of B.9 and G.30 compared with red light, but similar to white light. All the responses were cultivar dependent, and G.41 shoot growth was not positively affected by the above factors. However, increasing the culture time significantly improved the shoot growth of G.41. Ultrastructure and cell characteristics were examined from both abnormal and normal shoot apices. The cell anatomy of stunted shoots was similar to the cell anatomy associated with bud dormancy. Thus, we concluded that the appearance of stunted in vitro shoots of apple rootstocks were primarily caused by the bud dormancy, which may be overcome by chilling for a proper duration, culturing under proper light quality, and applying proper plant growth regulators (PGRs). Future research to promote shoot proliferation and elongation of apple rootstocks and other woody species with the similar limitations should be conducted on breaking in vitro shoots’ bud dormancy