Description

The four subtracks in this annotation correspond to replication timing categories for DNA synthesis. Replication is segregated into early specific (Early), mid specific (Mid), late specific (Late), and non-specific (PanS). The first three categories correspond to regions that replicated in a time point-specific manner; the latter category encompasses regions that replicated in a temporally non-specific manner.

Display Conventions and Configuration

This annotation follows the display conventions for composite tracks. To display only selected subtracks, uncheck the boxes next to the tracks you wish to hide.

Methods

The experimental strategy adopted to map this profile involved isolation of replication products from HeLa cells synchronized at the G1-S boundary by thymidine-aphidicolin double block. Cells released from the block were labeled with BrDu at every two-hour interval of S-phase and DNA was isolated from them. The heavy-light (H/L) DNA representing the pool of DNA replicated during each two-hour labeling period was separated from unlabeled DNA by double cesium chloride density gradient centrifugation. The purified H/L DNA was then hybridized to a high-density genome-tiling Affymetrix array comprised of all unique probes within the ENCODE regions.

The time of replication of 50% (TR50) of each microarray probe was calculated by accumulating the sum over the five time points and linearly interpolating the time when 50% was reached. Each probe was also classified as temporally specific or non-specific based on whether or not at least 50% of the accumulated signal appeared in a single time point.

The TR50 data was then analyzed within a 20 kb sliding window to classify regions as specific versus non-specific based on the ratio of specific to non-specific probes within the window. Specific regions were further classified as early, mid, or late replicating based on the average TR50 of specific probes within the window. The resulting regions form a non-overlapping segregation of the replication data into the four given categories of replication timing.

Verification

The replication experiments were completed for two biological sets in the HeLa-adherent cell line.

Credits

Data generation and analysis for this track were performed by the DNA replication group in the Dutta Lab at the University of Virginia: Neerja Karnani, Christopher Taylor, Hakkyun Kim, Louis Lim, Ankit Malhotra, Gabe Robins and Anindya Dutta.

Neerja Karnani and Christopher Taylor prepared the data for presentation in the UCSC Genome Browser.

References

Jeon, Y., Bekiranov, S., Karnani, N., Kapranov, P., Ghosh, S., MacAlpine, D., Lee, C., Hwang, D.S., Gingeras, T.R. and Dutta, A. Temporal profile of replication of human chromosomes. Proc Natl Acad Sci U S A 102(18), 6419-24 (2005).