Abstract
The most basic level of eukaryotic gene regulation is the presence or absence of nucleosomes on DNA regulatory elements. In an effort to elucidate in vivo nucleosome patterns, in vitro studies are frequently used. In vitro, short DNA fragments are more favorable for nucleosome formation, increasing the likelihood of nucleosome occupancy. This may in part result from the fact that nucleosomes prefer to form on the terminal ends of linear DNA. This phenomenon has the potential to bias in vitro reconstituted nucleosomes and skew results. If the ends of DNA fragments are known, the reads falling close to the ends are typically discarded. In this study we confirm the phenomenon of end bias of in vitro nucleosomes. We describe a method in which nearly identical libraries, with different known ends, are used to recover nucleosomes which form towards the terminal ends of fragmented DNA. Finally, we illustrate that although nucleosomes prefer to form on DNA ends, it does not appear to skew results or the interpretation thereof.
Highlights
Chromatin is the combination of DNA and DNA-associated proteins
End bias occurs near the ends of DNA fragments
These studies looking at end bias occurring in vitro, were based on low throughput data, did not quantify the amount of end biased nucleosome positioning, and did not analyze if this end bias affects down-stream analyses
Summary
Chromatin is the combination of DNA and DNA-associated proteins. A histone octamer, made up of eight histone proteins, serves as the first means of DNA compaction and organization [1]. End bias could only be analyzed for the RsaI and HincII invitrosome libraries, as they are the only invitrosome libraries where the DNA fragments used in the reconstitution had defined ends.
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