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Plant Biotechnology Reports

, Volume 3, Issue 3, pp 175–182 | Cite as

High frequency direct plant regeneration from leaf, internode, and root segments of Eastern Cottonwood (Populus deltoides)

  • Rakesh Yadav
  • Pooja Arora
  • Dharmendar Kumar
  • Dinesh Katyal
  • Neeraj Dilbaghi
  • Ashok Chaudhury
Original Article

Abstract

Simple, reproducible, high frequency, improved plant regeneration protocol in Eastern Cottonwood (Populus deltoides) clones, WIMCO199 and L34, has been reported. Initially, aseptic cultures established from axillary buds of nodal segments from mature plus trees on MS liquid medium supplemented with 0.25 mg l−1 KIN and 0.25 mg l−1 IAA. Nodal and internodal segments were found to be extra-prolific over shoot apices during course of aseptic culture establishment, while 0.25 mg l−1 KIN concentration played a stimulatory role in high frequency plant regeneration. Diverse explants, such as various leaf segments, internodes, and roots from in vitro raised cultures, were employed. Direct plant regeneration was at high frequency of 92% in internodes, 88% in leaf segments, and 43% in root segments. This led to the formation of multiple shoot clusters on established culture media with rapid proliferation rates. Many-fold enhanced shoot elongation and growth of the clusters could be achieved on liquid MS medium supplemented with borosilicate glass beads, which offer physical support for proliferating shoots leading to faster growth in comparison to semi-solid agar or direct liquid medium. SEM examination of initial cultures confirmed direct plant regeneration events without intervening calli. In vitro regenerated plants induced roots on half-strength MS medium with 0.15 mg l−1 IAA. Rooted 5- to 6-week-old in vitro regenerated plants were transferred into a transgenic greenhouse in pots containing 1:1 mixture of vermicompost and soil at 27 ± 2°C for hardening and acclimatization. 14- to 15-week-old well-established hardened plants were transplanted to the field and grown to maturity. The mature in vitro raised poplar trees exhibited a high survival rate of 85%; 4-year-old healthy trees attained an average height of 8 m and an average trunk diameter of 25 cm and have performed well under field conditions. The regeneration protocol presented here will be very useful for undertaking genetic manipulation, providing a value addition to Eastern Cottonwood propagation in future.

Keywords

Eastern Cottonwood Internodes Leaf Direct plant regeneration Populus deltoides Root segments 

Abbreviations

BAP

6-Benzylaminopurine

IAA

Indole-3-acetic acid

KIN

Kinetin

MS

Murashige and Skoog

SEM

Scanning electron microscopy

Notes

Acknowledgments

This work was supported by the Department of Biotechnology, Ministry of Science & Technology, Government of India, New Delhi as a research project to Prof. A. Chaudhury. Mr. Rakesh Yadav, Mr. Dharmendar Kumar and Mr. Dinesh Katyal duly acknowledge financial assistance in the form of JRF’s and Technical Assistant in Department of Biotechnology sponsored project. The authors wish to express their gratitude to Dr. H.C. Chaturvedi, Emeritus Scientist, NBRI (CSIR), Lucknow and Dr. Vibha Dhawan, Vice Chancellor, TERI University for providing plant material and rendering help in initial phase of work. The transgenic greenhouse, tissue culture facility were established under the FIST program, and financial assistance in form of color reproduction fee for manuscript figure was provided under BT-19 Project, Department of Science & Technology, Ministry of Science & Technology, Government of India, New Delhi.

References

  1. Bolstad PV, Libby WJ (1982) Comparison of radiata pine cutting of hedge and tree form origin after seven growing seasons. Silvae Genet 31:9–13Google Scholar
  2. Borchert R (1976) The concept of rejuvenility in woody plants. Acta Hortic 56:21–36Google Scholar
  3. Chaturvedi HC, Sharma AK, Agha BQ, Jain M, Sharma M (2004) Production of clonal trees of Populus deltoides through in vitro regeneration of shoots from leaf, stem and root explants and their field cultivation. Indian J Biotechnol 2:203–208Google Scholar
  4. Chaudhury A, Qu R (2000) Somatic embryogenesis and plant regeneration of turf type Bermudagrass: effect of 6-Benzyladenine in callus induction medium. Plant Cell Tissue Organ Cult 60:113–120CrossRefGoogle Scholar
  5. Coleman GD, Ernst SG (1989) In vitro shoot regeneration of Populus deltoides: effect of cytokinin and genotype. Plant Cell Rep 8:459–462CrossRefGoogle Scholar
  6. Coleman GD, Ernst SG (1990) Shoot induction competence and callus determination in Populus deltoides. Plant Sci 71:83–92CrossRefGoogle Scholar
  7. Greenwood MS (1987) Rejuvenation of forest trees. Plant Growth Regul 6:1–12CrossRefGoogle Scholar
  8. Hackett WP (1985) Juvenility, maturation, and rejuvenation in woody plants. Hortic Rev 7:109–155Google Scholar
  9. Halperin W (1973) The use of cultured tissue in studying developmental problems. Can J Bot 51:1801–1806CrossRefGoogle Scholar
  10. Hu WJ, Harding SA, Lung J, Popko JL, Ralph J, Stokke DD, Tsai CJ, Chiang VL (1999) Repression of lignin biosynthesis promotes cellulose accumulation and growth in transgenic trees. Nat Biotechnol 17:808–812PubMedCrossRefGoogle Scholar
  11. Jansson S, Douglas CJ (2007) Populus: a model system for plant biology. Annu Rev Plant Biol 58:435–458PubMedCrossRefGoogle Scholar
  12. Klee HJ, Horsch RB, Hinchee MA, Hein MB, Hoffman NL (1987) The effects of overproduction of two Agrobacterium tumefaciens T-DNA auxin biosynthetic gene products in transgenic petunia plants. Genes Dev 1:86–96CrossRefGoogle Scholar
  13. MacLeod K, Nowak J (1990) Glass beads as a solid matrix in in vitro study of the role of polyamines in cold hardiness of white clover. Plant Cell Tissue Org Cult 22:113–117CrossRefGoogle Scholar
  14. Michler CH, Bauer EO (1991) High frequency somatic embryogenesis from leaf tissue of Populus Spp. Plant Sci 77:111–118CrossRefGoogle Scholar
  15. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497CrossRefGoogle Scholar
  16. Son SH, Hall RB (1990) Plant regeneration capacity of callus derived from leaf, stem, and root segments of Populus alba L.× P. grandidantata Michx. Plant Cell Rep 9:344–347CrossRefGoogle Scholar
  17. Stoutemyer VT, Britt OK (1965) Tissue cultures of juvenile and adult specimens of ivy. Nature 199:397–398CrossRefGoogle Scholar
  18. Stoutemyer VT, Britt OK (1969) Growth and habituation in tissue cultures of English ivy, Hedera helix. Am J Bot 56(2):222–226CrossRefGoogle Scholar
  19. Thakur AK, Sharma DK (2006) High efficiency plant regeneration from leaf explants of male Himalayan poplar (P. ciliata wall). In Vitro Cell Dev Biol Plant 42:144–147CrossRefGoogle Scholar
  20. Thakur AK, Sharma S, Srivastava DK (2005) Plant regeneration and genetic transformation studies in petiole tissue of Himalayan poplar (Populus ciliata Wall.). Curr Sci 89:664–668Google Scholar
  21. Tsvetkov I, Hausman JF, Jouve L (2007) Thidiazuron-induced regeneration in root segments of white poplar (P. alba L.). Bulg J Agric Sci 13:623–626Google Scholar
  22. Vasil V, Vasil IK (1984) Preparation of cultured tissues for scanning electron microscopy. In: Vasil IK (ed) Cell culture and somatic cell genetics of plants. Laboratory procedures and their applications, vol 1. Academic Press, New York, USA, pp 738–743Google Scholar
  23. Winten LL (1968) Plantlet formation from aspen tissue culture. Science 160:1234–1235CrossRefGoogle Scholar
  24. Wolter KE (1968) Root and shoot initiation in aspen callus culture. Nature 219:503–510CrossRefGoogle Scholar

Copyright information

© Korean Society for Plant Biotechnology and Springer 2009

Authors and Affiliations

  • Rakesh Yadav
    • 1
  • Pooja Arora
    • 1
  • Dharmendar Kumar
    • 1
  • Dinesh Katyal
    • 1
  • Neeraj Dilbaghi
    • 1
  • Ashok Chaudhury
    • 1
  1. 1.Department of Bio and Nano TechnologyGuru Jambheshwar University of Science and TechnologyHisarIndia

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