EpiScope Nucleosome Preparation Kit
EpiScope Nucleosome Preparation Kit
Brand: Takara Bio.
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EpiScope Nucleosome Preparation Kit
This kit is designed for the preparation of nucleosomes from cultured mammalian cells. DNA can be extracted from the prepared nucleosome and analyzed using a real-time PCR assay or high-speed sequencing to map nucleosome positions.
Overview
- Enables detailed analysis of nucleosome positioning
- Prepared DNA consists primarily of mononucleosomes
Applications
- Preparation of nucleosomal DNA from 0.5–2 x 106 mammalian cultured cells
- Nucleosome positioning with regards to DNA methylation analysis
Genomic DNA regions occupied with nucleosomes are quantified or identified with downstream applications, such as qPCR, DNA sequencing, and microarray
Genomic DNA regions occupied with nucleosomes are quantified or identified with downstream applications, such as qPCR, DNA sequencing, and microarray.
Protocol overview
Protocol overview.
Analysis of nucleosomal DNA size distribution
Analysis of nucleosomal DNA size distribution. Nucleosomal DNA preparation from HeLa cells (1 x 106 cells) was performed with various amounts of micrococcal nuclease (0.02 , 0.05, and 0.2 ul). Nucleosomal DNA was purified with the NucleoSpin Extract II, yielding 50 ul of DNA solution, of which 5 ul was analyzed by agarose gel electrophoresis. For samples treated with lower amounts of micrococcal nuclease (0.02 ul, 0.05 ul), the electrophoretic results showed bands for mononucleosomal, dinucleosomal, and trinucleosomal DNA fragments. For samples treated with 0.2 ul of micrococcal nuclease, mostly mononucleosomes were evident. In addition, the higher the amount of micrococcal nuclease used, the shorter the size of the mononucleosome-derived DNA fragment observed. This result may reflect the trimming activity at both ends of the DNA (Clark et al. 2010).
Clark, D. J. Nucleosome positioning, nucleosome spacing and the nucleosome code. J. Biomol. Struct. Dyn. 27, 781–93 (2010).
Dutta, A. & Workman, J. L. Nucleosome Positioning: Multiple Mechanisms toward a Unifying Goal. Mol. Cell 48, 1–2 (2012).
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Genomic DNA regions occupied with nucleosomes are quantified or identified with downstream applications, such as qPCR, DNA sequencing, and microarray
Genomic DNA regions occupied with nucleosomes are quantified or identified with downstream applications, such as qPCR, DNA sequencing, and microarray.
Protocol overview
Protocol overview.
Analysis of nucleosomal DNA size distribution
Analysis of nucleosomal DNA size distribution. Nucleosomal DNA preparation from HeLa cells (1 x 106 cells) was performed with various amounts of micrococcal nuclease (0.02 , 0.05, and 0.2 ul). Nucleosomal DNA was purified with the NucleoSpin Extract II, yielding 50 ul of DNA solution, of which 5 ul was analyzed by agarose gel electrophoresis. For samples treated with lower amounts of micrococcal nuclease (0.02 ul, 0.05 ul), the electrophoretic results showed bands for mononucleosomal, dinucleosomal, and trinucleosomal DNA fragments. For samples treated with 0.2 ul of micrococcal nuclease, mostly mononucleosomes were evident. In addition, the higher the amount of micrococcal nuclease used, the shorter the size of the mononucleosome-derived DNA fragment observed. This result may reflect the trimming activity at both ends of the DNA (Clark et al. 2010).
