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Genomic and RNA Profiling Core Facility

Houston, Texas

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Genomic and RNA Profiling Core
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ChIP Sequencing Services

Illumina Chromatin Immunoprecipitation Sequencing (ChIP-Seq)

Researchers can expand the scale of their study to identify binding sites of DNA-associated proteins across the entire genome simultaneously using Illumina’s ChIP-Seq. This is useful in determining gene regulation by using information about CpG methylation, histone modifications, chromatin structure or DNA-protein interactions and allows the analysis of protein/nucleic acid interactions and binding events without being confined to set number of array features or candidate sequences. A library is prepared using the small fragments of DNA that were cross linked to the protein of interest, fractionated then enriched. Oligonucleotide adapters are added to each end of the fragments and clustered on a flow cell. The TruSeq ChIP sample Preparation Kit can multiplex up to 24 samples that can be run in a single lane.

The first step of library preparation by the core is a sample quality check. During sample quality check, the quantification of each sample by NanoDrop and Picogreen will be determined. Typically, the concentration of ChIP DNA samples will be too low to accurately read by NanoDrop. Multiple ChIP’d samples can be pooled together, up to 50ul, to obtain the minimum 5-10ng of ChIP DNA. A bioanalyzer chip is used to determine the size range of each sample. The majority of DNA fragments should fall between 100-400bp. Larger fragment sizes will be discarded during the size selection step of the library preparation protocol.

ChIP-Seq Input Amounts
GARP recommended Minimum ChIP'd DNA Input GARP recommended Minimum DNA Concentration
10 ng (5 ng)* 0.2 ng/μl (0.1 ng/μl)*
*Minimum Assay Requirements

*NOTE: Fold enrichment of the samples must be given to continue with library preparation. A 4-fold enrichment is required. ChIP-Seq libraries will be given to users to perform qPCR assay to confirm that a minimum 2-fold enrichment is maintained after library preparation.

Additional information:

- DNA shearing can be done using enzymatic methods, such as MNase or by physical shearing, such as Covaris, Bioruptor or tip probe sonicator. Get the Bioruptor User Guide.

- Carrier DNA used as a blocking agent during the immunoprecipitation process can serve as a template during library construction and will form clusters when sequencing.

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