Nucleic Acid Research Group 2013–2014 Study: Evaluating Library Synthesis Protocols for Sub-Nanogram ChIPSeq samples

摘要

Chromatin immunoprecipitation followed by sequencing the precipitated DNA (ChIP-Seq) is the state-of-the-art method to study protein-DNA interactions. ChIP-Seq allows identification of binding sites of proteins across the entire genome in an unbiased manner. One of the major limitations of currently available ChIP-Seq protocols is the necessity to isolate sufficient amounts of immune precipitated DNA for subsequent sequencing. The NARG 2013/14 study evaluated library preparation alternatives starting from one and two orders of magnitude DNA less than standard protocols. Library preparation kits from seven different commercial providers were utilized in this project. Some of these kits were intended for low input while other kits were used outside of specifications of the manufacturers. Aliquots of the same preparation of ChIPed DNA were processed using the standard protocol for 10 ng of input DNA and the evaluated library preparation alternatives for 1 ng and 100 pg of input DNA. Each library type was prepared at two different ABRF Member labs and sequencing was done on a single Illumina HiSeq flow cell.The results of low input compared to the standard protocol will be presented. The NARG 2013/14 study provides information how ChIP-Seq can be performed from 100 times less DNA than by standard methods with minimal compromising quality of results.