Tissue Culture Literation
| Plant latin name | Paeonia lactiflora Pallas |
| Literature code | Paeonia_lactiflora-Ref-3 |
| Reference | Wu HJ et al., New Zealand Journal of Crop and Horicultural Science 39: 271-278 (2011) |
| Summary | The underground buds of herbaceous peony (Paeonia lactiflora Pall.) ‘Zhong Sheng Fen’ were used to investigate the effects of indole-3-acetic acid (IAA) on culture initiation, benzyladenine (BA) on axillary shoot induction, and three auxins (IAA, indole-3-butyric acid [IBA] and 1-naphthyleneacetic acid [NAA]) on axillary shoot proliferation. In addition, methods to rejuvenate hyperhydric microshoots were established. Our results showed that the best initiation medium for ‘Zhong Sheng Fen’ was half-strength Murashige and Skoog (MS) medium supplemented with double the concentration of Ca2+ (880 mg L-1), 1 mg L-1 BA, 0.5 mg L-1 gibberellic acid (GA3) and 0.1 mg L-1 IAA. Axillary shoots were successfully induced by 2.0 mg L-1 BA. Several methods allowed the successful rejuvenation of hyperhydric microshoots, allowing them to develop normally. These included the addition of 3 g L-1 activated charcoal, the removal of ammonium nitrate from the medium, doubling the concentration of Ca2+ or eliminating BA from the medium. |
| Objectives | Induction of axillary shoots and exploration of methods to optimize the rejuvenation of hyperhydric microshoots |
| Materials | P. lactiflora ‘Zhong Sheng Fen’ from the Society of Forestry Experimental Base |
| Explant | The well-developed underground buds of P. lactiflora ‘Zhong Sheng Fen’ were collected in winter from the Society of Forestry Experimental Base and used as the explants. |
| Initial culture | Before inoculation, the underground buds were washed in tap water for 30 min. The outer scales were peeled off and buds were soaked in 75% ethanol for 30 s, immediately followed by 10 min sterilization with a dilute solution of HgCl2 (0.1% w/v). Plant material was rinsed five times (5 min/rinse) in autoclaved distilled water. All sterilized explants (i.e. underground buds) were placed on agar (0.7%) solidified medium. The base medium was half-strength (macro- and micronutrients) MS medium (Murashige and Skoog 1962) with doublestrength calcium chloride (Ca2+ ) supplemented with 30 g L-1 sugar. Buds were cultured in 100 ml Erlenmeyer flasks containing 30 ml agar solidified medium with one bud per flask. The effects of indole-3-acetic acid (IAA) (0.0, 0.1, 0.3 and 0.5 mg L-1) combined with 1 mg L-1 6-benzyladenine (BA) and 0.5 mg L-1 gibberellic acid (GA3) on culture initiation of ‘Zhong Sheng Fen’ were tested. The highest lateral shoot initiation (92.8%), number of lateral shoots (2.13) and shoot length (4.2 cm) and the lowest rate of yellowing shoot tips (17.43%) were observed in medium containing 0.1 mg L-1 IAA + 1 mg L-1 BA + 0.5 mg L-1 GA3. |
| Shoot multiplication | After initiation of clture, the effects of BA (0.5, 1.0, 1.5 and 2.0 mg L-1) combined with 0.5 mg L-1 kinetin (KT) on axillary shoot induction were tested. The induction of axillary shoots was favoured by a higher concentration of BA (2.0 mg L-1). The effects of different auxins (IAA, IBA, NAA) on axillary shoot proliferation were tested. The addition of an auxin to the medium caused a significant increase in the incidence of hyperhydricity compared with the control. The highest multiplication rate (2.40) and the lowest incidence of hyperhydricity (7.01%) were observed on auxin- free medium (0.2 mg L-1 BA + 0.1 mg L-1 GA3). Even a low concentration of BA (0.2 mgL-1) aggravated hyperhydric microshoots. In this state, no hyperhydric microshoots reversed to normal shoots and the proliferation rate of hyperhydric microshoots also decreased. Hyperhyfricity was strongly linked to BA, and even a low concentration of BA (0.2 mgL-1) did not benefit the growth of hyperhydric microshoots. Several methods allowed the successful rejuvenation of hyperhydric microshoots, allowing them to develop normally. These included the addition of 3 g L-1 activated charcoal, the removal of ammonium nitrate from the medium, doubling the concentration of Ca2+ or eliminating BA from the medium. |
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