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Samples were separated by SDS-PAGE (NuPage 412% Bis-Tris Gel, Life Technologies), run with BioRad Precision Plus Protein Kaleidoscope ladder, and transferred to a nitrocellulose membrane using an iBlot system (Life Technologies)

However, it is limited by its qualitative or semi-quantitative nature, relatively low specificity, the need for precise primer design, and challenges arising from uneven 5mC distribution (62)