Seyyed Mohammad Hossein Hayatolgheibi; Ali Akbar Mozafari
Abstract
Introduction: The major problem in apple well-known rootstocks is lack of protocols for fast propagation under in vitro condition. Nitric oxide (NO) has been received the great encouragement and more attention in the recent years for its key signaling role. Nitric oxide plays a vital role in the growth ...
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Introduction: The major problem in apple well-known rootstocks is lack of protocols for fast propagation under in vitro condition. Nitric oxide (NO) has been received the great encouragement and more attention in the recent years for its key signaling role. Nitric oxide plays a vital role in the growth and development of plants, including stimulating the seed germination and seedlings growth as well as delaying in the senescence process.
In previous studies, the application of sodium nitroprusside (SNP), as NO-releasing agent, in combination with different plant hormones under in vitro conditions showed that, The application of 30 μM SNP significantly increased shoot multiplication (9.4 shoots per explant) and the use of 100 μM SNP induced rhizogenesis (2.1 roots per explants) of apple micro cutting. Accordingly, the current study attempted to investigate the effects of SNP treatments in combination with NAA and BA on the regeneration of adventitious shoots and in combination with IBA and NAA on rhizogenesis of micro cuttings in MM111 and MM106 apple rootstocks, , under in vitro conditions.
Materials and Methods: The current study was conducted to investigate the effects of SNP alone and in combination with different types of growth regulators (IBA, NAA and BA) on the morpho-physiological characteristics of Malling Merton 111 (MM111) and Malling Merton 106 (MM106) micro cuttings under in vitro conditions. MM111 and MM106 that growth under in vitro conditions were already used with about 2.5 cm length as the plant's sources. This research was carried out in the frame of two separate experiments (proliferation and rhizogenesis). For the proliferation, the MS medium supplemented with different concentrations of SNP (0.0, 2.96, 5.98, 8.94, 11.91 and 14.90 mg L-1) used as treatments. For the rhizogenesis, the ½ MS medium supplemented with different concentrations of SNP (0, 7.45, 14.90, 22/35 and 57.80 mg L-1) alone and combined with 1 mg L-1 IBA and 0.01 mg L-1 NAA was used. In the first experiment, characteristics such as shoot length, number of shoots, total soluble proteins and carbohydrates content, peroxidase activity, carotenoids, chlorophyll a, chlorophyll b as well as total chlorophyll content were measured. In the rhizogenesis experiment, root length, fresh and dry weight of roots, as desirable characteristics, were measured. In both experiments, the treatments were arranged in a completely randomized factorial design with four replicates. Four and three explants were used in each replication for proliferation and rhizogenesis experiments, respectively.
Results and Discussion: In the proliferation experiment, the number of shoots under 5.98 mg L-1 SNP was significantly higher than other treatments. The experimental treatments did not have a significant effect on the shoots length. Since nitric oxide may play a role in cell division, so it participates in the regeneration of the lateral branches and caused their proliferation (11). The results showed that total chlorophyll and carbohydrate contents in MM106 rootstock were significantly higher than MM111. The highest total chlorophyll content was observed in 5.98 and 14.90 mg L-1 SNP treatments and the maximum soluble carbohydrates was obtained in 2.96 mg L-1 SNP treatment. Shoot regeneration under SNP treatments had a relatively high correlation with the amount of soluble proteins and carbohydrates. In the rhizogenesis experiment, the root length at 5.98, 11.91 and 14.90 mg L-1 SNP treatments were significantly different from other treatments. The lowest root number was observed in the absence of SNP. The previous literature indicated that NO induces the CYCD3:1 gene and caused the expression of the anti-CDK inhibitor KPP2 gene at the onset of the formation of peripheral lateral root, and the genetic regulators of auxin-dependent cell cycle is directly related to NO. Also, our results showed that root fresh weight under 5.98 and 14.90 mg L-1 SNP treatments was significantly higher than other treatments, and the highest root dry weight was obtained in 5.98 mg L-1 SNP in comparison to other treatments. Based on the results it may be assumed that presence of SNP causes changes in the level of plant hormones at different stages of development, which is probably resulted in starting metabolic processes for root development and dry matter accumulation. Each trait showed a more favorable result at a specific concentration of SNP. However, proliferation under 5.96 mg L-1 SNP first increased then reduced.
Conclusion: Application of SNP treatments had a positive effect on the measured traits e.g. shoot numbers, total soluble protein and carbohydrate contents, as well as fresh and dry weight of roots. In this experiment, the concentration of 5.98 mg L-1 SNP had the highest effect in term of shoot numbers, total soluble protein and carbohydrate contents, compared to other treatments. The apple rootstock MM106 showed the better performance to the plant growth regulators than MM111 rootstock. Overall, the present results indicated that SNP material, as a NO-releasing source, can physiologically be present in the plant in a way that can induce regeneration of plants and this potential depends on the genotype type.
