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A Hypomethylated population of Brassica rapa for forward and reverse Epi-genetics

Stephen Amoah1, Smita Kurup1, Carlos Marcelino Rodriguez Lopez3, Sue J Welham1, Stephen J Powers1, Clare J Hopkins14, Michael J Wilkinson3 and Graham J King12*

Author Affiliations

1 Rothamsted Research, Harpenden, Herts AL5 2JQ, UK

2 Current address: Southern Cross Plant Science, Southern Cross University, Lismore, NSW 2480, Australia

3 Plant Research Centre, School of Agriculture, Food and Wine, Faculty of Sciences, University of Adelaide, Waite Campus, PMB1, Glen Osmond, SA, 5064, Australia

4 Department of Pathology, The University of Melbourne, Melbourne, VIC, 3010, Australia

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BMC Plant Biology 2012, 12:193  doi:10.1186/1471-2229-12-193

Published: 20 October 2012

Additional files

Additional file 1:

Figure S1. Dosage response curve for seedling growth of B. rapa R-o-18 raised from seeds pre-treated with 5-AzaC. B. rapa seeds were treated with five different concentrations of 5-AzaC (0.01mM, 0.1mM, 0.5mM, 1.0mM and 1.5mM) and a control. A gradient of retarded growth was observed with increasing concentration. This was quantified by measuring the height of individual plants (Figure 1a). Figure S2. Effect of 5-AzaC treatments on flowering time in B. rapa R-o-18. Seeds were treated with five different concentrations of 5-AzaC (0.01mM, 0.1mM, 0.5mM, 1.0mM and 1.5mM) and a water control. Days to flowering, defined as the number of days that had lapsed since seeds were sown in soil to the day the first anthesis was observed on the plant. Data points in graph represent the mean number of days to flowering at the concentrations studied. Figure S3. Box-plot analysis of E2 seed weight resulting from 5-AzaC treatment. Seed were harvested from plants derived from seeds that had been treated with 5-AzaC. At lower concentrations of 5-AzaC some E1 plants had heavier seeds and others with lighter seeds. The box represents 50% of plants. The whiskers at the top and the bottom of the boxes represent the upper and the lower quartiles respectively. The horizontal line across the boxes indicates the median seed weight.

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Additional file 2:

Table S1. Primers used for MSAP analysis. Selective nucleotides are indicated as +XYZ in the primer code column. Enzyme column indicates the restriction enzyme site associate with each primer. Table S2. Epigenetic molecular diversity induced by 5-AzaC. Epigenetic diversity induced by 5-AzaC calculated using Analysis of Molecular Variance (AMOVA) inferred from the analysis of methylation-sensitive amplified polymorphism (MSAP) assays using primer combinations H2/E1 and H3/E3. Populations are ordered following their PhiPT values which indicate the epigenetic distance between each population restricted with HpaII and MspI (Populations highlighted in red presented significantly lower PhiPTs when compared to the original B. rapa line R-o-18). Prob indicates the probability of having a more extreme variance component and PhiPT than the observed values by chance alone. The Sum of Squares within population (SSWP) reflects intra-population diversity from the analysis of methylation-sensitive amplified polymorphism (MSAP) assays using the methylation sensitive restriction enzyme HpaII.

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Additional file 3:

Table S3. Genes identified as upregulated in Brassica rapa line E3 line BraRoAZ_12445e3 compared with the R-o-18 control line. Leaf RNA was hybridised against the GeneChip Brassica Exon 1.0 ST Array [3940]. Results were analysed using GeneSpring v.11.5, with GO analysis essentially following that described by Love et al. [40].

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