Abstract
Somaclones exhibiting variations with flower characteristics were recovered from the tissue-culture-derived plants of Doritaenopsis. Two molecular techniques, random amplified polymorphic DNA (RAPD) and methylation-sensitive amplification polymorphism (MSAP) analyses, were used to characterize the somaclones. RAPD analysis, using 100 randomly selected primers, failed to differentiate variants and normal plants, even though some primers (six out of 100 primers) exhibited 6–10 distinct banding patterns. However, MSAP analysis revealed the differences in the DNA methylation patterns in the normal and variant plants which were correlated with phenotypic variation. In all, 311, 337, 366, and 343 fragments were obtained with normal and V1, V2, and V3 variant plants, respectively; each representing recognition site cleaved by either or both of the isoshizomers were amplified using 12 combination of primers. A total of 36 (11.6%), 77 (22.9%), 73 (19.9%), and 47 (13.7%) sites were found to be methylated at cytosine in the genomes of normal and V1, V2, and V3 variant Doritaenopsis plants. This study demonstrates usefulness of MSAP to detect DNA methylation events in tissue cultured Doritaenopsis plants.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Arnholdt-Schmitt B. Rapid changes in amplification and methylation pattern of genomic DNA in cultured carrot root explants. Theor. Appl. Genet. 85-85: 793–800; 1993. doi:10.1007/BF00225021.
Arnholdt-Schmitt B.; Herterich S.; Neumann K. Physiological aspects of genome variability in tissue culture. I. Growth phase-dependent differential DNA methylation of carrot genome (Daucus carota L.) during primary culture. Theor. Appl. Genet. 91: 809–815; 1995.
Bajaj Y. P. S. Somaclonal variation—origin, induction, cryopreservation, and implication in plant breeding. In: Bajaj Y. P. S. (ed) Biotechnology in agriculture and forestry, vol.2: Somaclonal variation in crop improvement I. Springer, Berlin, pp 3–48; 1990.
Baurens P.; Bonnot F.; Bienvenu D.; Causse S.; Legavre T. Using SD-AFLP and MSAP to assess CCGG methylation in the banana genome. Plant. Mol. Biol. Rep. 21: 339–348; 2003. doi:10.1007/BF02772583.
Chen W. H.; Chen T. M.; Fu Y. M.; Hsich R. M.; Chen W. S. Studies on somaclonal variation in Phalaenopsis. Plant Cell Rep. 18: 7–13; 1998. doi:10.1007/s002990050523.
Finnegan E. J.; Genger R. K.; Peacock W. J.; Dennis E. S. DNA methylation in plants. Annu. Rev. Plant Physiol. Plant Mol. Biol. 49: 223–247; 1998. doi:10.1146/annurev.arplant.49.1.223.
Kaeppler S. M.; Kaeppler H.; Rhee Y. Epigenetic aspects of somaclonal variation in plants. Plant Mol. Biol. 43: 179–188; 2000. doi:10.1023/A:1006423110134.
Li X.; Xu M.; Korban S. S. DNA methylation profiles differ between field- and in vitro-grown leaves of apple. J. Plant Physiol. 159: 1129–1134; 2002.
LoSchiavo F.; Pitto L.; Giuliano G.; Torti G.; NutiRonchi V.; Vergara R.; Orselli S.; Terzi M. DNA methylation of embryogenic carrot cells and its variation as caused by mutation, differentiation, hormones and hypomethylating drugs. Theor. Appl. Genet. 77: 325–331; 1989. doi:10.1007/BF00305823.
Park S. Y.; Murthy H. N.; Paek K. Y. Rapid propagation of Phalaenopsis from floral stalk derived leaves. In Vitro Cell Dev. Biol- Plant 38: 168–172; 2002. doi:10.1079/IVP2001274.
Park S. Y.; Murthy H. N.; Paek K. Y. Protocorm-like body induction and subsequent plant regeneration from root tip cultures of Doritaenopsis. Plant Sci. 164: 919–923; 2003. doi:10.1016/S0168-9452(03)00019-0.
Peraza-Echeverria S.; Herrera-Valencia V. A.; James-Kay A. Detection of DNA methylation changes in micropropagated plants using methylation-sensitive amplification polymorphism (MSAP). Plant Sci. 161: 359–367; 2001. doi:10.1016/S0168-9452(01)00421-6.
Peredo E. L.; Rvilla M. A.; Arroyo-Garcia R. Assessment of genetic and epigenetic variation in hop plants regenerated from sequential subcultures of organogenic calli. J. Plant Physiol. 163: 1071–1079; 2006. doi:10.1016/j.jplph.2005.09.010.
Reina-Lopez G. R.; Simpson J.; Ruiz-Herrera J. Differences in DNA methylation patterns are detectable during dimorphic transition of fungi by amplification of restriction polymorphism. Mol. Gen. Genet. 253: 703–710; 1997. doi:10.1007/s004380050374.
Tokuhara K. Somaclonal variation of flowers in micropropagated plants through flower stalk bud culture. In: Ichihashi S.; Nagata H. (eds) Proc Nagoya Int Orchid Show 92. Nagoyapp, Japan, pp 317–319; 1992.
Xiong L. Z.; Xu C. G.; Saghal Maroof M. A.; Zhang Q. Patterns of cytosine methylation in an elite rice hybrid and its parental lines, detected by a methylation-sensitive amplification polymorphism technique. Mol. Gen. Genet. 261: 439–446; 1999. doi:10.1007/s004380050986.
Acknowledgements
This work was supported by Korean Science and Engineering Foundation. One of the authors (H.N. Murthy) is thankful to Korean Federation of Science and Technology Societies for the award of Brain Pool Fellowship.
Author information
Authors and Affiliations
Corresponding author
Additional information
Editor: C.N. Stewart
Rights and permissions
About this article
Cite this article
Park, S.Y., Murthy, H.N., Chakrabarthy, D. et al. Detection of epigenetic variation in tissue-culture-derived plants of Doritaenopsis by methylation-sensitive amplification polymorphism (MSAP) analysis. In Vitro Cell.Dev.Biol.-Plant 45, 104–108 (2009). https://doi.org/10.1007/s11627-008-9166-6
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11627-008-9166-6