Morphological and Anatomical Investigations on Old Home x Farmingdale 333 / Pyrus elaeagrifolia in Vitro Micrografts

Abstract

In this study, the possibility of creation of a specific pear rootstock combination by micrografting was investigated. Two genotypes; ‘Old Home x Farmingdale 333’ (Pyrus communis L.) which is a clonal semi dwarf pear rootstock resistant to fireblight and pear decline, and Pyrus elaeagrifolia known as tolerant to Fe-chlorosis, salinity and drought stresses were used to obtain a clonal, semi dwarf and tolerant to the stresses pear rootstock combination. In vitro P. elaeagrifolia seedlings were used as rootstock, and in vitro shoots of ‘OH x F 333’ were used as semi dwarf interstock. Cleft grafting was applied in micrografts. Grafted seedlings were cultured on Murashige and Skoog basal medium with ½ strength of macronutrients for 8 weeks. Cultures were incubated under white fluorescent light, 16 h per day. However, complete darkness was applied for 1 or 2 weeks at the beginning of incubation for the darkness treatments. Graft take success in the control treatment was significantly higher (97.9%) than darkness treatments (90.5% and 82.5%, respectively). Ultrastructural observations with transmission electron microscope revealed that dividing cambial initials reached to 2-3 layers and new xylem and phloem elements distinctly differentiated in transversal sections of the graft union after 8 weeks of micrografts in the control and dark treatments. The results indicated that a successful graft union formation occurred. To the best of our knowledge, this is the first report of in vitro micrografting on Pyrus elaeagrifolia Pallas.

Keywords

• Micrografting; Graft union; Interstock; Dark treatment

Morphological and Anatomical Investigations on in Vitro Micrografts of OHxF 333 / Pyrus elaeagrifolia Interstock / Rootstock Combination in Pears

Abstract

In this study, possibility of creating a specific interstock / rootstock combination to obtain a clonal, semi dwarf, composite pear rootstock tolerant to various stresses by micrografting was investigated. In vitro shoots of ‘Old Home x Farmingdale 333’ (Pyrus communis L.) which is a clonal semi dwarf pear rootstock resistant to fireblight and pear decline was used as interstock, and in vitro Pyrus elaeagrifolia Pallas seedlings known as tolerant to Fe-chlorosis, salinity and drought stresses was used as rootstock. Cleft grafting was applied in micrografts. Grafted seedlings were cultured on Murashige and Skoog basal medium with ½ strength of macronutrients for 8 weeks under white fluorescent light for 16 h day. Cultures, except the control, received complete darkness either 1 or 2 weeks at the beginning of incubation. Graft take success in the control treatment was significantly higher (97.9%) than darkness treatments of 1 or 2 weeks (90.5% and 82.5%, respectively). Ultrastructural observations with transmission electron microscope revealed that dividing cambial initials reached to 2-3 layers, and new xylem and phloem elements distinctly differentiated in transverse sections of the graft union 8 weeks after micrografting in the control and darkness treatments. The results indicated a successful graft union formation.

Keywords

• Micrografting; Graft union; Interstock; Darkness treatment

References

1. Abousalim A & Mantell S H (1992). Micrografting of pistachio (Pistacia vera L. cv. Mateur). Plant Cell, Tissue and Organ Culture 29: 231-234
2. Amiri M A (2006). In vitro techniques to study the shoot-tip grafting of Prunus avium L. (cherry) var. Seeyahe Mashad. Journal of Food Agriculture and Environment 4: 151-154
3. Barba M, Cupidi A, Loreti S, Faggioli F & Martino L (1995). In vitro micrografting: a technique to eliminate peach latent mosaic viroid from peach. Acta Horticulturae 386: 531-535
4. Bell R L (1990). Pears (Pyrus). In: Moore J N & Ballington J R (Eds), Genetic Resources of Temperate Fruit and Nut Crops, International Society for Horticultural Science, Wageningen, pp. 655-696
5. Bisognin C, Ciccotti A, Salvadori A, Moser M, Grando M S & Jarausch W (2008). In vitro screening for resistance to apple proliferation in Malus species. Plant Pathology 57: 1163-1171
6. Can C, Ozaslan M, Toremen H, Sarpkaya K & Iskender E (2006). In vitro micrografting of pistachio, Pistacia vera L. var. Siirt, on wild pistachio rootstocks. Journal of Molecular Cell Biology 5: 25-31
7. Conejero A, Romero C, Cunill M, Mestre M A, MartínezCalvo J, Badenes M L & Llácer G (2013). In vitro shoot-tip grafting for safe Prunus budwood exchange. Scientia Horticulturae 150: 365-370
8. Dobranszki J & Silva J A T (2010). Micropropagation of apple - A review. Biotechnology Advances 28: 462488

9. Edriss M H & Burger D W (1984). Micro-grafting shoottip culture of Citrus on three trifoliolate rootstocks. Scientia Horticulturae 23: 255-259
10. Elivar D E & Dumanoğlu H (1999). Ayaş (Ankara) koşullarında elma, armut ve ayvada bir yaşlı fidan üretiminde ilkbahar sürgün ve sonbahar durgun göz aşılarının karşılaştırılması. Tarım Bilimleri Dergisi 5(2): 58-64
11. Espen L, Cocucci M & Sacchi G A (2005). Differentiation and functional connection of vascular elements in compatible and incompatible pear / quince internode micrografts. Tree Physiology 25: 1419-1425
12. Estrada-Luna A A, Lopez-Peralta C & Cardenas-Soriano E (2002). In vitro micrografting and the histology of graft union formation of selected species of pricly pear cactus (Opuntia spp.). Scientia Horticulturae 92: 317-327
13. Faggioli F, Martino L & Barba M (1997). In vitro micrografting of Pyrus communis shoot tips. Advances in Horticultural Science 11: 25-29
14. Ghorbel A, Chatibi A, Thaminy, S & Kchouk M L (1999). Micrografting of almond Prunus dulcis (Miller) D.A. Webb.) cv. Achak. Acta Horticulturae 470: 429-433
15. Han Y Y, Woo J H, Sim Y G, Choi K B, Choi B S & Yeo S K (1998). Studies on the plug seedling of rose rootstock (Rosa multiflora ‘Hort No. 1’). RDA Journal of Horticulture Science 40: 82-88
16. Hartmann H T, Kester D E, Davies F T & Geneve R L (2011). Plant Propagation: Principle and Practices. Prentice-Hall, London
17. Hassanen S A (2013). In vitro grafting of pear (Pyrus spp.). World Applied Sciences Journal 21: 705-709
18. Huang S C & Millican D F (1980). In vitro micrografting of apple shoot tips. HortScience 15: 741-743
19. Isıkalan C, Namli S, Akbas F & Ak B E (2011). Micrografting of almond (Amygdalus communis) cultivar ‘Nonpareil’. Australian Journal of Crop Science 5: 61-65
20. Jarausch W, Lansac M, Bliot C & Dosbat F (1999). Phytoplasma transmission by in vitro graft inoculation as a basis for a preliminary screening method for resistance in fruit trees. Plant Pathology 48: 283-287
21. Jonard R (1986). Micrografting and its applications to tree improvement. In: Bajaj Y P S (Ed). Biotechnology in Agriculture and Forestry, Springer, Berlin, pp. 31-48
22. Ke S, Cai Q & Skirvin R M (1993). Micrografting speeds growth and fruiting of protoplast-derived clones of kiwifruit (Actinidia deliciosa). Journal of Horticultural Science 68: 837-840
23. Lane W D, Bhagwat B, Armstrong J D & Wahlgren S (2003). Apple micrografting protocol to establish transgenic clones on field ready rootstock. HortTechnology 13: 641-646
24. Lombard P B & Westwood M N (1987). Pear rootstocks. In: Rom R C & Carlson R F (Eds). Rootstocks for Fruit Crops, A Wiley-Interscience Publication, John Wiley and Sons, Inc., New York, pp. 145-183
25. Luft J H (1961). Improvements in epoxyresin embedding methods. The Journal of Biophysical and Biochemical Cytology 9: 409-414
26. Ma F, Guo C, Liu Y, Li M, Ma T, Mei L & Hsiao A (1996). In vitro shoot-apex grafting of mulberry (Morus alba L.). HortScience 31: 460-462
27. Matsumoto K, Tamura F, Chun J P & Tanabe K (2006). Native Mediterranean Pyrus rootstock, P. amygdaliformis and P. elaeagrifolia, present higher tolerance to salinity stress compared with Asian Natives. Journal of the Japanese Society for Horticultural Science 75: 450-457
28. Monteuuis O (1994). Effect of technique and darkness on the success of meristem micrografting of Picea abies. Silvae Genetica 43: 91-95
29. Monteuuis O (1996). In vitro shoot apex micrografting of mature Acacia mangium. Agroforestry Systems 34: 231-217
30. Monteuuis O (2012). In vitro grafting of woody species. Propagation of Ornamental Plants 12: 11-24
31. Murashige T & Skoog F (1962). A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiologia Plantarum 15: 473-497
32. Murashige T, Bitters W P, Rangan T S, Nauer E M, Roistacher C N & Holliday B P (1972). A technique of shoot apex grafting and its utilization towards recovering virus free Citrus clones. HortScience 7: 118-119
33. Musacchi S, Ganino T, Grassi S & Fabbri A (2004). Grafting in vitro shoots on acclimating plants: morphological and anatomical aspects of compatible and incompatible pear / quince combinations. Acta Horticulturae 658: 559-563
34. Nas M N & Read P E (2003). Simultaneous micrografting, rooting and acclimatization of micropropagated American chestnut, grapevine and hybrid hazelnut. European Journal of Horticultural Science 68: 234237
35. Nava R, Garcia A V, Marin C & Villegas Z (2011). Clonal propagation of elite Carica papaya L. plants using in vitro and in vivo micrografting. Interciencia 36: 517523
36. Navarro L, Roistacher C N & Murashige T (1975). Improvment of shoot tip grafting in vitro for virus free Citrus. Journal of the American Society for Horticultural Science 100: 471-479
37. Nunes J C O, Abreu M F, Dantas A C M, Pereira A J & Pedrotti E L (2005). Morphological characterization in apple micrografts. Revista Brasileira de Fruticultura 27: 80-83
38. Obeidy A A & Smith M A L (1991). A versatile new tactic for fruit tree micrografting. HortTechnology 1: 91-95
39. Onay A, Pirinc V, Adıyaman F, Isıkalan C, Tilkat E & Basaran D (2003). In vivo and in vitro micrografting of pistachio, Pistacia vera L. cv. “Siirt”. Turkish Journal of Biology 27: 95-100
40. Onay A, Pirinc V, Yıldırım H & Basaran D (2004). In vitro micrografting of mature pistachio (Pistacia vera var. Siirt). Plant Cell, Tissue and Organ Culture 77: 215219
41. Ozden-Tokatlı Y, Akdemir H, Tilkat E & Onay A (2010). Current status and conservation of Pistacia germplasm. Biotechnology Advances 28: 130-141
42. Ozzambak E & Schmidt H (1991). In vitro and in vivo micrografting of cherry (Prunus avium L.). Gartenbauwissenschaft 56: 221-223
43. Parthasarathy V A, Nagaraju V & Rahman S A S (1997). In vitro grafting of Citrus reticulata Blanco. Folia Horticulturae 9: 87-90
44. Sato J E (1967). A modifred method for load staining of thin sections. Journal of Electron Microscopy 16: 133 Sharma S, Singh B, Rani G, Zaidi A A, Halan V K, Nagpal A K & Virk G S (2008). In vitro production of Indian citrus ringspot virus (ICRSV) free Kinnow plants employing thermotherapy coupled with shoot 279 Volk G M, Bonnart R, Krueger R & Lee R (2012). Cryopreservation of citrus shoot tips using micrografting for recovery. CryoLetters 33: 418-426
45. Yıldırım H, Onay A & Suzerer V (2010). Micrografting of almond (Prunus dulcis Mill.) cultivars “Ferragnes” and “Ferraduel”. Scientia Horticulturae 125: 361367
46. Yin H, Yan B, Sun J, Jia P, Zhang Z, Yan X, Chai J, Ren Z, Zheng G & Liu H 2012. Graft-union development: a delicate process that involves cell– cell communication between scion and stock for local auxin accumulation. Journal of Experimental Botany 63: 4219-4232