بررسی تاثیر نیتریک اکسید (NO) بر پرآوری و ریشه‌زایی ریزقلمه پایه‌های سیب MM111 و MM106 در شرایط درون‌شیشه‌ای

نوع مقاله : مقالات پژوهشی

نویسندگان

دانشگاه کردستان

چکیده

پژوهش حاضر به منظور بررسی تاثیر سدیم نیتروپروساید (SNP) و تنظیم­کننده­های رشد بر ویژگی­های مورفو-فیزیولوژیکی گیاهچه­های پرآوری شده و ریشه­زایی آنها انجام گردید. این مطالعه شامل دو آزمایش فاکتوریل در قالب طرح کاملاً تصادفی با چهار تکرار بود. به منظور پرآوری میکروشاخه‌ها از محیط­کشت پایه­MS­ حاوی تنظیم­کننده­های رشد (یک میلی­گرم در­ لیتر بنزیل­آدنین (BA) به همراه 01/0 میلی­گرم در لیتر نفتالین‌استیک­اسید(NAA) ) و SNP در شش سطح شامل صفر، 96/2، 98/5، 94/8، 91/11، 90/14میلی­گرم در لیتر بر روی دو پایه سیب MM106 وMM111 استفاده شد. در آزمایش ریشه­زایی به بررسی اثر SNP (صفر، 45/7، 91/11،90/14، 35/22­و80/57 میلی­گرم­در ­لیتر) به تنهایی و در ترکیب با یک میلی­گرم در لیتر IBA و 01/0 میلی­گرم بر لیتر NAA روی محیط­کشت پایه MS­½پرداختیم. پس از 60  روز شاخص­های رشدی ساقه و ریشه شامل طول شاخه و ریشه، تعداد شاخه و ریشه، وزن تر و خشک ریشه، پروتئین­های­محلول­کل، فعالیت­آنزیم پراکسیداز، کربوهیدرات­محلول­کل، کاروتنوئید و کلروفیل­a، b و­ کل مورد اندازه­گیری قرار گرفتند. نتایج نشان داد که روند تغییرات شاخه­زایی تحت تاثیر تیمارهایSNP همسویی نسبتا بالایی با مقدار پروتئین­های محلول و کربوهیدارت داشت بطوری که با افزایش میزان SNP به 98/5 میلی­گرم در لیتر، مقدار هر سه پارامتر اندازه­گیری شده افزایش و سپس کاهش یافت. همچنین روند تغییرات مقدار کاروتنوئید و کلروفیل گیاهچه­ها با تغییرات مقدار SNP همبستگی نداشت. بیشترین تعداد ریشه در تیمارهای­91/11 و­ 35/22 میلی­گرم در لیتر SNP­به همراه یک­ میلی­گرم در لیترIBA  و 01/0 میلی­گرم در لیتر NAAبدست آمد، در حالیکه بیشترین طول ریشه در تیمارهای 45/7، 91/11 و 90/14 میلی­گرم در لیترSNP  حاصل شد. لذا غلظت­های مختلفSNP  و نیز ترکیب­SNP  با تنظیم کننده­های می­توانند نقش موثری روی اندام­زایی پایه­های سیب داشته باشند.

کلیدواژه‌ها


عنوان مقاله [English]

The Study of Nitric Oxide (NO) Effect on Proliferation and Rhizogenesis of the MM106 and MM111 Apple Rootstocks Micro Cutting under In vitro Conditions

نویسندگان [English]

  • S. M. H. Hayatolgheibi
  • ALI AKBAR MOZAFARI
چکیده [English]

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.

کلیدواژه‌ها [English]

  • morphology
  • Physiology
  • Proliferation
  • Rhizogenesis
1- Booij-James I.S., Edelman M. and Mattoo A.K. 2009. Nitric oxide donor-mediated inhibition of phosphorylation shows that light-mediated degradation of photosystem II D1 protein and phosphorylation are not tightly linked. Planta, 229:1347-1352.
2- Bradford M.M. 1979. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72:248-254.
3- Chen Y.H., Chao Y.Y., Hsu Y.Y., Hong C.Y. and Kao C.H. 2012. Hemeoxygenase is involved in nitric oxide- and auxin-induced lateral root formation in rice. Plant Cell Reports, 31:1085–1091.
4- Chohan A., Parmar U. and Raina S.K. 2012. Effect of sodium nitroprusside on morphological characters under chilling stress in chickpea (Cicer arietinum L.). Journal of Environmental Biology, 33:695-698.
5- Correa-Aragunde N., Graziano M. and Lamattina L. 2004. Nitric oxide plays a central role in determining lateral root development in tomato. Planta, 218:900–905.
6- Correa-Aragunde N., Graziano M., Chevalier C. and Lamattina L. 2006. Nitric oxide modulates the expression of cell cycle regulatory genes during lateral root formation in tomato. Journal of Experimental Botany, 57:581–588.
7- Cui X.M., Zhang Y.K., Wu X.B. and Liu C.S. 2010. The investigation of the alleviated effect of copper toxicity by exogenous nitric oxide in tomato plants. Plant Soil Environment, 56:274–281.
8- Gao Z., Lin Y., Wang X., Wei M., Yang F. and Shi Q. 2014. Sodium nitroprusside (SNP) alleviates the oxidative stress induced by NaHCO3 and protects chloroplast from damage in cucumber. African Journal of Biotechnology, 11:6974-6982.
9- Gibson S.I. 2005. Control of plant development and gene expression by sugar signaling. Current Opinion in Plant Biology, 8:93-102.
10- Gouvea C.M.C.P., Souza J.F., Magalhas A.C.N. and Martins I.S. 1997. NO releasing substances that induce growth elongation in maize root segments. Plant Growth Regulator, 21:183–187.
11- Han X., Yang H., Duan K., Zhang X., Zhao H., You S. and Jiang Q. 2009. Sodium nitroprusside promotes multiplication and regeneration of Malus hupehensis in vitro plantlets. Plant Cell, Tissue and Organ Culture, 96:29–34.
12- Hemeda H.M. and Kelin B.P. 1990. Effects of naturally occurring antioxidants on peroxidase activity of vegetables extracts. Journal of Food Science, 55:184-185.
13- Huang A.X. and She X.P. 2003. Effect of SNP on Rhizogenesis of hypocotyls cutting from mung bean seedling. Acta Botanica Boreali-Occidentalia Sinica, 23:2196–2199. (in Chinese with English abstract).
14- Irigoyen J.J., Emerich D.W. and Sanchez-Diaz M. 1992. Water stress induced changes in concentrations of proline and total soluble sugars in nodulated alfalfa (Medicago sativa) plants. Journal of Plant Physiology, 84:55-60.
15- Jhanji S., Setia R.C., Kaur N., Kaur P. and Setia N. 2012. Role of nitric oxide in cadmium-induced stress on growth, photosynthetic components and yield of Brassica napus L. Journal of Environmental Biology, 33:1027-1032.
16- Kalra C. and Babbar S.B. 2010. Nitric oxide promotes in vitro organogenesis in Linum usitatissimum L. Plant Cell, Tissue and Organ Culture, 103:353–359.
17- Kolberz Z., Bartha B. and Erdei L. 2008. Exogenous auxin-induced NO synthesis is nitrate reductase-associated in Arabidopsis thaliana root primordial. Journal of Plant Physiology, 65:967–975.
18- Lichtenthaler H.K. and Buschmann C. 2001. Extraction of photosynthetic tissues: chlorophylls and carotenoids. Food Analytical Chemistry, F4. 2.1-F4. 2.6.
19- Molnar Z., Virag E. and Ördög V. 2011. Natural substances in tissue culture media of higher plants. Acta Biologica Szegediensis, 55:123-127.
20- Murashige T. and Skoog F. 1962. A revised medium for rapid growth and bio-assays with tobacco tissue cultures. Physiology of Plant, 15:473-497.
21- Neill S.J. and Hancock J.T. 2003. Nitric oxide signaling in plants. New Phytology, 159:11–35.
22- Pagnussat G.C., Lanteri M.L. and Lamattina L. 2003. Nitric oxide and cyclic GMP are messengers in the indole acetic acid-induced adventitious Rhizogenesis process. Plant Physiology, 132:1241–1248.
23- Pagnussat G.C., Lanteri M.L., Lombardo M.C. and Lamattina L. 2004. Nitric oxide mediated the indole acetic acid induction activation of a mitogen-activated protein kinase cascade involved in adventitious root development. Plant Physiology, 135:279–286.
24- Prochazkova D., Haisel D., Wilhelmova N., Pavlikova D. and Szakova J. 2013. Effects of exogenous nitric oxide on photosynthesis. Photosynthetica, 51(4):483-489.
25- Sarropoulou V., Dimassi-Theriou K. and Therios I. 2014. Ιn vitro plant regeneration from leaf explants of the cherry rootstocks CAB-6P, Gisela 6, and MxM 14 using sodium nitroprusside. In Vitro Cellular and Developmental Biology - Plant, 50:226-234.
26- Sarvajeet S.G. and Narendra T. 2010. Reactive oxygen species and antioxidant machinery in abiotic stress tolerance in crop plants. Plant Physiology and Biochemistry, 48:909–930.
27- Tan C.B., Chin C.F. and Alderson P. 2013. Effects of sodium nitroprusside on shoot multiplication and regeneration of Vanilla planifolia Andrews. In vitro Cellular and Developmental Biology - Plant, 49:626-630.
28- Tavallali V. and Rahemi M. 2007. Effects of Rootstock on Nutrient Acquisition by Leaf, Kernel and Quality of Pistachio (Pistacia vera L.). American-Eurasian Journal of Agricultural and Environmental Science, 2:240-246.
29- Xu, J., Yin H., Wang W., Mi Q. and Liu, X. 2009. Effects of sodium nitroprusside on callus induction and shoot regeneration in micropropagated Dioscorea opposita. Plant Growth Regulation. 59:279–285.