Quantifying DNA Replication Dynamics
Corresponding Organization : Tokyo University of Pharmacy and Life Sciences
Variable analysis
- Incubation time of the extract sample with Cy5-dUTP
- Replication activity (measured by Cy5 signal)
- Total genomic DNA (measured by SYBR Gold signal)
- Extract sample volume (10 μl)
- Cy5-dUTP concentration (2 μM)
- Stop buffer composition (5-mM EDTA, 200-mM NaCl, 0.5% SDS, 20-mM Tris-HCl, pH 8.0)
- Proteinase K concentration (200 μg/ml)
- RNaseA concentration (10 μg/ml)
- Incubation time for proteinase K and RNaseA treatment (2 h at 37 °C)
- Agarose gel concentration (0.8% for neutral condition, 1% for denaturing condition)
- Gel electrophoresis buffer (Tris-acetate EDTA, TAE)
- Staining method (SYBR Gold)
Annotations
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