Description
The EpiNext™ cTIP (CUT&Tag In-Place)-Sequencing Kit is a complete set of reagents required for carrying out a successful cTIP-Seq starting from mammalian cells. The kit is designed to enrich a protein (histone or strong binding transcription factor)- specific DNA complex from low input cells/chromatin and to prepare a library for next-generation sequencing using Illumina platforms such as Illumina Genome Analyzer II, HiSeq, and MiSeq systems. The innovative working principle, optimized protocol, and components of the kit allow for the capture of targeted protein/DNA complexes with minimized non-specific background levels and for the construction of both non-barcoded (singleplexed) and barcoded (multiplexed) libraries quickly in order to map target protein-DNA interaction regions with less bias and at a high resolution. The kit has the following advantages and features:
- High enrichment: Uses a unique nucleic acid cleavage enzyme mix, which has low sequence bias, to simultaneously fragment chromatin and cleave/remove any DNA sequences in both ends of the target protein/DNA complex without affecting DNA occupied by the target protein. Thus, the true target protein-enriched regions can be reliably identified and high-resolution mapping can be achieved.
- Low input materials: Robust sonication-free fragmentation, unbound DNA cleavage, and immunocapture are all processed in the same single-tube with on-beads ligation. This method allows for using in both cells and tissues and allows for maximal degradation protection of the target protein with minimal sample loss. As a result, the input cell amount can be as few as 500 cells or chromatin amount can be as low as 50 ng.
- In place tagging: The on-beads ligation (in place) of DNA adapters to chromatin prior to DNA purification results in an increase in DNA fragment size, allowing too short fragments (ex: < 70 bps) bound to transcription factors (TF) to be purified for library construction.
- Minimal background: Cleavage of unbound DNA sequences in the two (2) end of the target protein/DNA complex in situ enables the minimized immunocapture/sequencing background, allowing data analysis with <10 million reads.
- Fast, streamlined procedure: The procedure from cells to library DNA is less than 5 hours.
- Highly convenient: The kit contains all required components for each step of the CUT&Tag in-place sequencing, which are sufficient for both protein/DNA capture and captured DNA library preparation, thereby allowing the cTIP (CUT&Tag In-Place)-Sequencing Kit to be the most convenient with reliable and consistent results.