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Tissue culture-induced locus-specific alteration in DNA methylation and its correlation with genetic variation in Codonopsis lanceolata Benth. et Hook. f

  • Genetics and Genomics
  • Published:
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Abstract

We have reported recently that tissue culture induced a high level of genetic variation at the primary nucleotide sequence in regenerants of medicinal plant Codonopsis lanceolata. It is not known, however, whether epigenetic variation in the form of alteration in DNA methylation also occurred in these plants. Here, we investigated possible alterations in level and pattern of cytosine methylation at the CCGG sites in the same set of regenerants relative to the donor plant, by the MSAP method employing a pair of isoschizomers, HpaII and MspI, which recognize the same restriction site but are differentially sensitive to cytosine methylation at the CCGG sites. A total of 1,674 MSAP profiles were resolved using 39 primer combinations. Of these, 177 (10.5%) profiles were polymorphic among the regenerants and/or between the regenerant(s) and the donor plant, in EcoRI + HpaII or EcoRI + MspI digest but not in both, indicating alteration in cytosine methylation patterns of specific loci, though their estimated total level of methylation remained more or less the same as the donor plant. Gel blot analysis validated most of the variant MSAP profiles as bona fide alteration in methylation patterns. Correlation analysis between the MSAP data and the previously reported ISSR and RAPD data revealed significant correlations, suggesting their possible intrinsic interrelatedness. Thirty-seven typical variant MSAP profiles were isolated and sequenced, of which 5 showed significant homology to known-function genes, 2 to chloroplast sequences, whilst the rest 30 did not find a match in the database.

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Abbreviations

MSAP:

Methylation-sensitive amplified polymorphism

RAPD:

Randomly amplified polymorphic DNA

ISSR:

Inter-simple sequence repeat

CTAB:

Cetyl trimethylammonium bromide

UPGMA:

Unweighted pair-group method using arithmetic averages

PCA:

Principal coordinate analysis

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Acknowledgments

This study was supported by the Program for Changjiang Scholars and Innovative Research Team (PCSIRT) in University (#IRT0519) and the National Natural Science Foundation of China (30225003, 30430060). We are grateful to two anonymous reviewers for their valuable suggestions to improve the manuscript.

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Authors and Affiliations

  1. Laboratory of Plant Molecular Epigenetics, Institute of Genetics and Cytology, Northeast Normal University, Changchun, 130024, China

    W. L. Guo, R. Wu, H. Y. Wang, L. Gong & Bao Liu

  2. Key Laboratory for Applied Statistics of MOE, Northeast Normal University, Changchun, 130024, China

    H. Y. Wang & Bao Liu

  3. School of Life Science and Engineering, Qiqihar University, Qiqihar, 161006, China

    Y. F. Zhang, X. M. Liu & Z. H. Zhang

Authors
  1. W. L. Guo
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  2. R. Wu
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  3. Y. F. Zhang
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  4. X. M. Liu
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  5. H. Y. Wang
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  6. L. Gong
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  8. Bao Liu
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Corresponding author

Correspondence to Bao Liu.

Additional information

Communicated by H.S. Judelson.

W. L. Guo and R. Wu contributed equally to this work.

Electronic supplementary material

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299_2007_320_MOESM1_ESM.doc

Supplementary Table 1 Performance of selected primer pairs for MSAP analysis in Codonopsis lanceolata regenerants (Reg. 1 to 10) and their donor plant (NB: number of bands, PB: number of polymorphic bands, PPB: percentage of polymorphic bands) (DOC 101 KB)

299_2007_320_MOESM2_ESM.doc

Supplementary Table 2 Relationships among 10 Codonopsis lanceolata regenerants and the donor plant based on Dice similarity matrices derived from data for MSAP (a), ISSR (b) and RAPD (c) (DOC 108 KB)

299_2007_320_MOESM3_ESM.doc

Supplementary Table 3 Sequence analysis of variant MSAP profiles isolated from regenerants of Codonopsis lanceolata (DOC 119 KB)

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Guo, W.L., Wu, R., Zhang, Y.F. et al. Tissue culture-induced locus-specific alteration in DNA methylation and its correlation with genetic variation in Codonopsis lanceolata Benth. et Hook. f. Plant Cell Rep 26, 1297–1307 (2007). https://doi.org/10.1007/s00299-007-0320-0

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