Abstract
An efficient in vitro multiplication system via multiple shoot bud induction and regeneration has been developed in Chlorophytum arundinaceum using shoot crown explants. Optimum regeneration frequency (87%) and desirable organogenetic response in the form of de novo organized multiple shoot buds without an intervening callus phase was obtained on Murashige and Skoog's (MS) minimal organics medium containing 3% sucrose (w/v) supplemented with 4×10−6 M Kn and 2×10−6 MIBA. Axenic secondary explants with multiple shoot buds on subculturing elicited best response with 1×10−5 M Kinetin (Kn) and 5×10−6 M indole-3-butyric acid (IBA) giving rise to an average of 18.74 shoots per culture with mean shoot length of 7.6 cm ± 1.73. Varying molar ratios of either Kn/IBA or Kn/NAA revealed statistically significant differences in the regeneration frequencies among the phytohormone treatments. It was observed that the shoot bud differentiation and regeneration was influenced by the molar ratios of cytokinins/auxin rather than their relative concentrations. Healthy regenerated shoots were rooted in half strength MS basal medium containing 3% sucrose (w/v) supplemented with 5×10−6 M IBA. Following simple hardening procedures, rooted plantlets, were transferred to soil-sand (1:1; v/v) with more than 90% success. Genetic fidelity was assessed using random amplified polymorphic DNA (RAPD), karyotype analysis and meiotic behaviour of in vitro and in vivo plants. Five arbitrary decamers displayed same banding profile within all the micropropagated plants and in vivo explant donor. The cytological and molecular analysis complemented and compared well and showed no genomic alterations in the plants regenerated through shoot bud differentiation. High multiplication frequency, molecular, cytological and phenotypic stability ensures the efficacy of the protocol developed for the production and conservation of this important endangered medicinal herb.
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Abbreviations
- BAP:
-
6-benzylaminopurine
- IBA:
-
Indole-3-butyric acid
- NAA:
-
α-napthaleneacetic acid
- IAA:
-
Indole-3-acetic acid
- 2,4-D:
-
2,4-dichlorophenoxyacetic acid
- Kn:
-
Kinetin (6-furfuryl-amino purine)
- RAPD:
-
Random Amplified Polymorphic DNA
- PCR:
-
Polymerase Chain reaction
- MS:
-
Murashige and Skoog's medium (1962)
References
Agretious TK, Martin KP, Hariharan M (1996) In vitro colonal multiplication of Alpinia calcarata Rose. Phytomorphology 46:133–138
Battaglia E (1955) Chromosome morphology and terminology. Caryologia 8:179–187
Bhat SR, Chandel KPS, Malik SK (1995) Plant regeneration from various explants of cultivated Piper species. Plant Cell Rep 14:398–402
Bohanec B, Jakse M, Ihan A, Javornik B (1995) Studies of gynogenesis in onion (Allium cepa). Plant Sci 104:215–224
Brown PTH, Lange FD, Kranz E, Lorz H (1993) Analysis of single protoplast and regenerated plants by PCR and RAPD technology. Mol Gen Genet 237:311–317
Cecchini E, Natah L, Cavallini A, Durante M (1992) DNA variation in regenerated plants of Pea. Theor Appl Genet 84:874–879
Chopra RN, Nayar SL, Chopra IC (1956) Glossary of Indian medicinal plants. CSIR, New Delhi, p 62
Cullis CA, Cleary W (1986) DNA variation in Flax tissue culture. Can J Genet Cytol 28:247–251
Doyle JJ, Doyle JL (1990) Isolation of plant DNA from fresh tissue. Focus 12:13–15
Dunstan DI, Thorpe TA (1986) Plant regeneration and genetic variability. In: Vasil IK (ed) Cell culture and somatic cell genetics of plants, vol 3. Academic Press, Orlando, pp 223–241
Ezura H, Oasawa K (1994) Ploidy of somatic embryos and the ability to regenerate plantlets in melon (Cucumis melo L.). Plant Cell Rep 14:107–111
Fourré JL, Berger P, Niquet L, André P (1997) Somatic embryogenesis and somaclonal variation in Norway spruce: morphogenetic, cytogenetic and molecular approaches. Theor Appl Genet 94:159–169
Gomez KA, Gomez AA (1984) Statistical procedures for agriculture research. John Wiley & Sons, New York, pp 7–83
Gupta PK, Varshney RK (1999) Molecular markers for genetic fidelity during micropropagation and conservation. Curr Sci 76:1308–1310
Isabel N, Tremblay L, Michaud M, Tremblay FM, Bousquet J (1993) RAPDs as an aid to evaluate the genetic integrity of somatic embryogenesis derived populations of Picea mariana. Theor Appl Genet 86:81–87
La cour LF (1941) Stain Technol 16:169
McLean KS, Lawrence GW, Reichert NA (1992) Callus induction and adventitious organogenesis of kenef (Hibiscus cannabinus L.)
Murashige T (1977) Plant cell and organ culture as horticultural practices. Acta Hortic 78:17–30
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497
Nag KK, Johri BM (1971) Morphogenetic studies on endosperm of some parasitic angiosperms. Phytomorphology 21:202–218
Narasimham KRSL, Ravuru BK (2003) A note on some endangered medicinal plants as NTFPs of Eastern Ghats, Andhra Pradesh. EPTRI-ENVIS Newsletter, vol 9: 2. EPTRI, Hyderabad, pp 11–12
Purohit D, Dave A (1996) Micropropagation of Sterculia urens Roxb.—an endangered tree species. Plant Cell Rep 15:704–706
Rani V, Parida A, Raina SN (1995) Random amplified polymorphic DNA (RAPD) markers for genetic analysis in micropropagated plants of Populus deltoids Marsh. Plant Cell Rep 14:459–462
Rani V, Raina SN (1998) Genetic analysis of enhanced axillary branching derived Eucalyptus tereticornis Smith and E. camaldulensis Dehn. plants. Plant Cell Rep 17:236–242
Skoog F (1971) Aspects of growth factor interactions in morphogenesis of tobacco tissue culture. In: Les cultures de Tissus de Plantes. Colloq Int C.N.R.S., Paris. No. 193, pp 115–135
Stebbins Jr GL (1950) Variation and evolution in plants. Oxford University Press, Oxford
Sunnichen VG, Shivanna KR (1998) Micropropagation of Eremostachys superba—an endangered endemic species from India. Curr Sci 74:699–702
The Wealth of India—a dictionary of Indian raw material and industrial products (2000) First supplement series (Raw material) National Institute of Science Communication, CSIR, New Delhi, vol 1: A-Ci, p 247
Tandon M, Shukla YN (1993) A bibenzyle xyloside from Chlorophytum arundinaceum. Phytochemistry 32:1624–1625
Tandon M, Shukla YN, Thakur RS (1992) 4, hydroxy-8,11-oxidoheneicosanol and other constituents from Chlorophytum arundinaceum roots. Phytochemistry 31:2525–2526
Vasil IK (1985) In: Henke R, Hughes K, Constanin M, Hollaender A (eds) Tissue culture in forestry and agriculture. Plenum Press, New York, pp 31–47
Verma S, Magotra R, Koul AK (2002) In vitro multiplication of Eremurus persicus Boiss. (Liliaceae)—an endangered species. Phytomorphology 52:315–321
Went FW (1938) Specific factors other than auxin affecting growth and root formation. Plant Physiol 13:55–80
Wochok ZS (1981)The role of tissue culture in preserving threatened and endangered plant species. Biol Conserv 20:83–89
Zahim-Al AM, Ford-Lloyed BV, Newbury HJ (1999) Detection of somaclonal variation in garlic (Allium sativum L.) using RAPD and cytological analysis. Plant Cell Reports 18:473–477
Acknowledgements
The authors thank Dr. G.N. Qazi, Director, Regional Research Laboratory (CSIR), Jammu, for useful discussion. We are also indebted to Dr. S. Pal, an Agronomist at RRLJ for marvelous cooperation during the course of present study. We also acknowledge the funding from GBPIHED, Almora, Uttranchal, India
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Communicated by P. P. Kumar
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Lattoo, S.K., Bamotra, S., Sapru Dhar, R. et al. Rapid plant regeneration and analysis of genetic fidelity of in vitro derived plants of Chlorophytum arundinaceum Baker—an endangered medicinal herb. Plant Cell Rep 25, 499–506 (2006). https://doi.org/10.1007/s00299-005-0103-4
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DOI: https://doi.org/10.1007/s00299-005-0103-4