Automated ChIPmentation procedure on limited biological material of the human blood fluke Schistosoma mansoni

Abstract

In living cells, the genetic information stored in the DNA sequence is always associated with chromosomal and extra-chromosomal epigenetic information. Chromatin is formed by the DNA and associated proteins, in particular histones. Covalent histone modifications are important bearers of epigenetic information and as such have been increasingly studied since about the year 2000. One of the principal techniques to gather information about the association between DNA and modified histones is chromatin immunoprecipitation (ChIP), also combined with massive sequencing (ChIP-Seq). Automated ChIPmentation procedure is a convenient alternative to native chromatin immunoprecipitation (N-ChIP). It is now routinely used for ChIP-Seq in many model species, using in general roughly 106 cells per experiment. Such high cell numbers are sometimes difficult to produce. Using the human parasite Schistosoma mansoni, whose production requires sacrificing animals and should therefore be kept to a minimum, we show here that automated ChIPmentation is suitable for limited biological material. We define the operational limit as ≥20,000 Schistosoma cells. We also present a streamlined protocol for the preparation of ChIP input libraries.

Heterochromatin protein 1 (HP1) ofSchistosoma mansoni: non-canonical chromatin landscape and fitness effects

“…Antibody titration was done as described in 16 with 0, 2, 4, 8 and 16μl antibody to obtain the saturating quantity and finally 8 μL of antibody HP1 AbCam (ab109028, lot GR38873387336-9) were used for each reaction. Input libraries were generated as described earlier and optimal number of library amplification cycles was determined by qPCR as described in the same protocol 26 .…”