Adventitious shoot regeneration from leaf and stem explants of Salix matsudana Koidz

Currently, the leaf and stem as explants for rapid propagation in vitro remained unknown in Salix matsudana. Multiple shoots were regenerated from leaf and stem (stem apex and stem with axil). The optimum medium for callus regeneration and shoots induction from leaf was on WPM medium containing 6.0 mg/l zeatin (Z) and 0.75 mg/l NAA. The optimum medium of shoot induction from stem apex was on MS medium supplemented with 8.0 mg/l Z and 1.0 mg/l NAA, and from stem with axil was on MS with 8.0 mg/l Z and 0.5 mg/l NAA. Rooting of regenerated shoots was obtained on the same medium supplemented with 1.0 mg/l activated charcoal.

Sodium Nitroprusside Stimulates Micropropagation and TDZ Induces Adventitious Shoots Regeneration in Red Flesh Apple Malus niedzwetzkyana Koehne Dieck ex

The aim of the present investigation was to optimize protocols for micropropagation and adventitious shoot regeneration from leaf explants of two wild ecotypes of red flesh apple Malus niedzwetzkyana for future breeding programs. At the proliferation stage, different concentrations of sodium nitroprusside (SNP) and triacontanol (TRIA) were compared. To optimize shoot regeneration from leaf explants, interactive effects of 1-phenyl-3-(1,2,3-thidiazol-5-yl)-urea – thidiazuron (TDZ), indole-3-butyric acid (IBA) and two explant types were investigated. At rooting stage, the effect of exposure time of microshoots to darkness and exposure time to different concentrations of IBA and α-naphthalene acetic acid (NAA) were compared. The results showed that SNP affected the growth rate significantly and the maximum multiplication rates per explant (9.6 in the first ecotype and 8.8 in the second) were produced in the Quoirin and Lepoivre medium containing 17 SNP µM, in addition to 4 µm 6-benzylaminopurine (BAP) and 3 µm gibberellic acid (GA3). IBA and TDZ affected the adventitious shoot regeneration from leaf explants significantly, the highest number of regenerated shoots (18.3 per explant) was obtained from the basal section of the leaves cultured on the medium containing 2 μM IBA and 15 μM TDZ. At rooting stage, the maximum rooting (88.6%) was obtained in the result of one weak exposure to darkness on medium containing 3 μM IBA.

Rejuvenation of chicory and lettuce plants following phase change in tissue culture

Background A frequent problem associated with the tissue culture of Compositae species such as chicory (Cichorium intybus L.) and lettuce (Lactuca sativa L.) is the premature bolting to in vitro flowering of regenerated plants. Plants exhibiting such phase changes have poor survival and poor seed set upon transfer from tissue culture to greenhouse conditions. This can result in the loss of valuable plant lines following applications of cell and tissue culture for genetic manipulation. Results This study demonstrates that chicory and lettuce plants exhibiting stable in vitro flowering can be rejuvenated by a further cycle of adventitious shoot regeneration from cauline leaves. The resulting rejuvenated plants exhibit substantially improved performance following transfer to greenhouse conditions, with increased frequency of plant survival, a doubling of the frequency of plants that flowered, and substantially increased seed production. Conclusion As soon as in vitro flowering is observed in unique highly-valued chicory and lettuce lines, a further cycle of adventitious shoot regeneration from cauline leaves should be implemented to induce rejuvenation. This re-establishes a juvenile phase accompanied by in vitro rosette formation, resulting in substantially improved survival, flowering and seed set in a greenhouse, thereby ensuring the recovery of future generations from lines genetically manipulated in cell and tissue culture.

Reinvigoration of diploid strawberry (Fragaria vesca) during adventitious shoot regeneration

Diploid strawberry (Fragaria vesca ‘Baiguo’) is a model plant for studying functional genomics in Rosaceae. Adventitious shoot regeneration is essential for functional genomics by Agrobacterium tumefaciens-mediated transformation. An efficient shoot regeneration method using diploid strawberry leaf explants was conducted on 1/2MS + 1/2B5 medium that contained 2.0 mg L−1 TDZ over 14 days of dark culture; this induced the maximum percentage of shoot regeneration (96.44 ± 1.60%) and the highest number of shoots per explant (23.46 ± 2.14) after 11 weeks of culture. The explants considerably enlarged after 12 days; then, turned greenish brown after 30 days, yellowish brown after 36 days, and completely brown and necrotic after 48 days. Large numbers of adventitious shoots were produced from 48 to 66 days, and the shoots elongated from 66 to 78 days; this represents a critical period of reinvigoration, which included 30 days for leaf explant chlorosis, 36 days for adventitious shoot appearance, and 48 days for generation of numerous shoots. During the reinvigoration process, higher expressions of the hormone synthesis-related genes Ciszog1, CKX2, CKX3, CKX7, YUC2, YUC6, YUC10, YUC9, and GA2ox were detected from 30 to 48 days. Our results indicate that these genes may regulate reinvigoration of shoot regeneration.

Adventitious Shoot Regeneration and Agrobacterium tumefaciens-mediated Transient Transformation of Almond × Peach Hybrid Rootstock ‘Hansen 536’

‘Hansen 536’ (Prunus dulcis × Prunus persica) is an important commercial rootstock for peach and almond. However, susceptibility to wet soil and bacterial canker has limited its use primarily to areas with less annual rainfall. Genetic engineering techniques offer an attractive approach to improve effectively the current problems with this cultivar. To develop an efficient shoot regeneration system from leaf explants, 10 culture media containing Murashige and Skoog (MS) or woody plant medium (WPM) supplemented with different plant growth regulators were evaluated, and adventitious shoot regeneration occurred at frequencies ranging from 0% to 36.1%. Optimal regeneration with a frequency of 32.3% to 36.1% occurred with WPM medium containing 8.88 µm 6-benzylamino-purine (BAP) and 0.98 to 3.94 µm indole-3-butyric acid (IBA). The regenerated shoots had a high rooting ability, and 80% of the in vitro shoots tested rooted and survived after being transplanted to substrate directly. Transient transformation showed an efficient delivery of the β-glucuronidase (GUS) reporter gene (gusA) using all three Agrobacterium tumefaciens strains tested with a concentration of OD600 0.5 to 1.0 for 4 days of cocultivation. The protocols described provide a foundation for further studies to improve shoot regeneration and stable transformation of the important peach and almond rootstock ‘Hansen 536’.