Abstract
Post-bisulfite adaptor tagging (PBAT) is a concept that enables the preparation of an efficient sequencing library from bisulfite-treated DNA, and it also means the protocol implemented the concept. Although the previous PBAT or rPBAT was sensitive enough for single-cell methylome analysis, the protocol had several drawbacks owing to the repeated random priming reactions. To resolve these problems, we developed a unique single-strand DNA ligation technique, termed TACS ligation, and established a new protocol called tPBAT. With tPBAT, the data quality improved, with a longer insert and higher map** rate than that obtained with rPBAT. In addition, paired-end sequencing and indexing were supported by the default. In this chapter, the tPBAT protocol is introduced, and a thorough description of its application to small samples is provided.
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Acknowledgments
This work was supported by JSPS KAKENHI through grants to F.M. (20H03243) and T.I. (17H06305) and AMED Platform Project for Supporting Drug Discovery and Life Science Research (Basis for Supporting Innovative Drug Discovery and Life Science Research (BINDS)) through a grant to T.I. (Grant Number JP20am0101103).
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Miura, F., Shibata, Y., Miura, M., Ito, T. (2023). Post-bisulfite Adaptor Tagging Based on an ssDNA Ligation Technique (tPBAT). In: Hatada, I., Horii, T. (eds) Epigenomics. Methods in Molecular Biology, vol 2577. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2724-2_2
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DOI: https://doi.org/10.1007/978-1-0716-2724-2_2
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