ASSESSMENT OF DNA METHYLATION CHANGES IN TISSUE CULTURE OF Brassica napus

摘要

Plant tissue culture, as a fundamental technique for genetic engineering, has great potential of epigenetic variation, of which DNA methylation is well known of importance to genome activity. We assessed DNA me-thylation level of explants during tissue culture of Brassica napus (cv. Yangyou 9), using high-performance liquid chromatography (HPLC) assisted quantification. By detecting methylation levels in hypocotyls cultured in mediums with different concentrations of hormones, we found dissected tissue cultured with 0.1 mg/L 2,4-D and 1.0 mg/L 6-BA, presented the lowest methylation level and highest induction rate of callus (91.0%). Different time point of cultured explants also showed obvious methylation variations, explants cultured after 6 and 21days exhibited methylation ratios of 4.33 and 8.07%, respectively. Whereas, the methylation ratio raised to 38.7% after 30 days cultivation, indicating that methylation level of hypocotyls ranged during tissue culture. Moreover, we observed that the methylation level in callus is the highest during regeneration of rape-seed, following the regenerated plantlets and hypocotyls. This paper indicated the function of hormones and differentiation of callus is relevant to the methylation levels during tissueculture.