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Analysis of somaclonal variation in transgenic and regenerated plants of Arabidopsis thaliana using methylation related metAFLP and TMD markers

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We provide evidence that nucleotide sequence and methylation status changes occur in the Arabidopsis genome during in vitro tissue culture at a frequency high enough to represent an important source of variation.

Abstract

Somaclonal variation is a general consequence of the tissue culture process that has to be analyzed specifically when regenerated plants are obtained in any plant species. Currently, there are few studies about the variability comprising sequence changes and methylation status at the DNA level, generated by the culture of A. thaliana cells and tissues. In this work, two types of highly reproducible molecular markers, modified methylation sensitive AFLP (metAFLP) and transposon methylation display (TMD) have been used for the first time in this species to analyze the nucleotide and cytosine methylation changes induced by transformation and tissue culture protocols. We found significantly higher average methylation values (7.5%) in regenerated and transgenic plants when compared to values obtained from seed derived plants (3.2%) and that the main component of the somaclonal variation present in Arabidopsis clonal plants is genetic rather than epigenetic. However, we have found that the Arabidopsis regenerated and transgenic plants had a higher number of non-fully methylated sites flanking transposable elements than the control plants, and therefore, their mobilization can be facilitated. These data provide further evidence that changes in nucleotide sequence and methylation status occur in the Arabidopsis genome during in vitro tissue culture frequently enough to be an important source of variation in this species.

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Acknowledgements

This work was supported by the Spanish DGICYT grant AGL2066-14249-C02-02 and by the Junta de Castilla y León grant LE052A06.

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  1. Área de Genética, Facultad de Ciencias Biológicas y Ambientales, Universidad de León, 24071, León, Spain

    Carlos J. Coronel, Ana I. González, María L. Ruiz & Carlos Polanco

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  1. Carlos J. Coronel
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  2. Ana I. González
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  3. María L. Ruiz
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  4. Carlos Polanco
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Correspondence to Carlos Polanco.

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Communicated by Tarek Hewezi.

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299_2017_2217_MOESM1_ESM.pptx

Supplementary Figure 1. Grouped column scatter graph depicting the number of different types of metAFLP markers detected in each plant analyzed corresponding to the CCGN and CCWG primer clusters. Red horizontal lines represent mean values and blue horizontal lines represent standard deviations. 208R0 and 211R0: regenerated cell lines; 261T0 and 291T0: transformed cell lines; R0s: grouped regenerated lines; T0s: grouped transformed lines; R0s+T0s: grouped cell lines (PPTX 428 KB)

299_2017_2217_MOESM2_ESM.pptx

Supplementary Figure 2. Grouped column scatter graph depicting the number of different types of TMD markers detected in each plant analyzed associated to copia transposable elements. Red horizontal lines represent mean values and blue horizontal lines represent standard deviations. 208R0 and 211R0: regenerated cell lines; 261T0 and 291T0: transformed cell lines; R0s: grouped regenerated lines; T0s: grouped transformed lines; R0s+T0s: grouped cell lines (PPTX 164 KB)

299_2017_2217_MOESM3_ESM.pptx

Supplementary Figure 3. Grouped column scatter graph depicting the number of different types of TMD markers detected in each plant analyzed associated to TRIM transposable elements. Red horizontal lines represent mean values and blue horizontal lines represent standard deviations. 208R0 and 211R0: regenerated cell lines; 261T0 and 291T0: transformed cell lines; R0s: grouped regenerated lines; T0s: grouped transformed lines; R0s+T0s: grouped cell lines (PPTX 190 KB)

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Coronel, C.J., González, A.I., Ruiz, M.L. et al. Analysis of somaclonal variation in transgenic and regenerated plants of Arabidopsis thaliana using methylation related metAFLP and TMD markers. Plant Cell Rep 37, 137–152 (2018). https://doi.org/10.1007/s00299-017-2217-x

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