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Yeda R&D Co. Ltd
Abstract ID: 1726
Barcoding is performed directly on the total cellular chromatin, thus avoiding the low-input enzymatic reactions occurring in existing ChIP. Importantly, this enables multiple chromatin-barcoded samples to be pooled for ChIP
Barcoding is performed directly on the total cellular chromatin, thus avoiding the low-input enzymatic reactions occurring in existing ChIP. Importantly, this enables multiple chromatin-barcoded samples to be pooled for ChIP in the same well, which further reduce initial input requirements and increasing cross-sample reproducibility. The centrifugation steps were minimized by fixing the cells before sorting and sonication. Next, the sheared chromatin is immobilized on magnetic beads coated with antibody to H3 and then indexed. The indexed chromatin is released from the H3 beads and pooled with chromatin from other samples. Finally, ChIP is performed with the desired antibody, and a single chromatin barcoded pool can be divided among multiple ChIP for profiling various chromatin modification.
The novel method was implemented in the research of Cellular differentiation, a process which is highly reliant on gene regulation and orchestrated by the chromatin landscape. These inconsistencies can be resolved by observing the chromatin dynamics during intermediate differentiation stages. Hematopoiesis provides a well-defined system for studying cellular differentiation. Importantly, dramatic chromatin reorganizations during hematopoiesis are critical for early hematopoiesis decisions. Today, due to the low cell numbers of the hematopoietic progenitors and additional technical limitations of working with in vivo samples, attaining dynamic chromatin maps of hematopoiesis are practically impossible. To overcome these limitations researchers from Dr. Ido Amit's lab at the Weizmann Institute developed iChIP. In addition to true small-cell numbers, iChIP offers better sensitivity, reproducible results and it is amenable to high-throughput experiments.
Indexing-first chromatin immunoprecipitation (iChIP) allows profiling of chromatin state in small cell numbers and in in vivo studies that could not be approached with conventional
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Last Updated May 2015