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Abstract
BackgroundIn mammals, DNA methylation at the 5-position of cytosine is the most essential epigenetic modification. Changes in the level of genome-wide DNA methylation (also known as overall DNA methylation) are associated with alterations in gene expression, thereby contributing to the phenotypic and physiological diversity. Current technologies for detecting overall DNA methylation either suffer from low sensitivity or require sophisticated equipment. Studies on domestic animals are hampered by the lack of complete and annotated genomic information.ResultsHere we report a rapid slot blot method using methyl-CpG binding protein (MBD1) to exam the level of overall DNA methylation in pigs and chickens. Using this rapid approach, we determined the methylation status in various DNA samples of a Chinese indigenous (Erhualian) and a Western (Large White) breed of pigs. We also chose day 18 embryos (E18) and newly hatched chicks (D1) of a Chinese indigenous chicken breed (Wen’s yellow-feathered broiler chicken) for genome-wide DNA methylation analysis. The results revealed tissue- and breed-specific differences, as well as age-dependent variations, in the level of overall DNA methylation.ConclusionThe results showed that the slot blot assay is a sensitive, highly specific and convenient method for semi-quantitative estimation of overall DNA methylation with no species specificity. This method does not require sophisticated equipment, such as high performance liquid chromatography (HPLC), or expensive technologies like sequencing, thus providing a useful tool for overall DNA methylation studies on domestic animals.
Highlights
In mammals, DNA methylation at the 5-position of cytosine is the most essential epigenetic modification
Sensitivity of the slot blot assay We examined the sensitivity of methyl-CpG-binding by incubating recombinant MBD1 protein with membranes carrying different quantities of genomic DNA
Genomic DNA extracted from the muscle of D1 chicken was used, and the result suggested that the slot blot routinely allowed detection of genomic DNA to a sensitivity of 0.5 μg (Figure 2)
Summary
DNA methylation at the 5-position of cytosine is the most essential epigenetic modification. Impairment of methylation can damage assorted aspects of gene regulation, such as X chromosome inactivation [1], genomic imprinting [4] and retroviral silencing [5]. Environmental factors, such as ionizing radiation [6] and hormone exposure [7], could influence the overall level of DNA methylation, which results in altered molecular pathways and increased risk of diseases. Analysis of genome-wide DNA methylation could provide an overview that may be missed in studies limited to specific genes of interest
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