Understanding the genome involves more than sequencing DNA. Epigenetic changes, including protein-DNA interactions, play a crucial role in regulating genome function and biology.  Chromatin immunoprecipitation (ChIP) is the method of choice for capturing protein-DNA interaction sites, such as transcription factor and histone binding sites. Nevertheless, analyzing the material from ChIP assays is a labor intensive, costly, qualitative and piecemeal exercise, limiting its use for high-throughput, quantitative, genome-wide applications. Helicos’s high throughput, single molecule sequencing technology lets you perform ChIP experiments like never before – giving you virtually bias-free, quantitative profiles of protein-DNA interactions in hundreds of samples for a very low cost per sample.

The Advantages
Helicos Single Molecule ChIP-Seq, an amplification-free assay with millions of reads per sample, provides you with:

• The ability to perform ChIP-Seq on hundreds of samples in a cost-effective manner
• The resolution to find changes not detectable on array-based platforms or amplification-based next generation sequencers
• The precision to ensure that your results stay the same from run-to-run and from prep-to-prep. The quantitative accuracy only possible with an amplification- and ligation-free approach
• The ability to analyze a variety of attomole-level DNA samples such as samples obtained from Methyl-Seq assays.

The application of the Helicos® Genetic Analysis System to ChIP-Seq has been described in recent publications:

Chromatin profiling by directly sequencing small quantities of immunoprecipitated DNA. Nat Methods (2010), 7, 47-49.

Mapping the regulon of Vibrio cholerae ferric uptake regulator expands its known network of gene regulation. Proc Natl Acad Sci U S A. 2011 Jul 26;108(30):12467-72. Epub 2011 Jul 12.

Genome-wide mapping of 5-hydroxymethylcytosine in embryonic stem cells. Nature. 2011 May 19;473(7347):394-7. Epub 2011 May 8.

The Evidence: Amplification-free ChIP-Seq on an Embryonic Stem Cell population
No other platform lets you analyze ChIP-Seq material directly, without amplification. ChIP assays typically provide no more than low nanogram quantities of DNA; amounts that are too low for direct use in ligation- and solid-phase PCR-based methods. Using a modified version of the standard Helicos DNA Sample Preparation methodology, scientists at Helicos were able to process minute amounts of DNA obtained from a collaborator’s ChIP experiment on a stem cell population, and elucidated protein-DNA binding events throughout the human genome (see Figure 1).

Figure 1. A ChIP DNA sample from a stem cell population and the corresponding input DNA sample were both processed without amplification using a modified version of the Helicos DNA Sample Preparation methodology described in Figure 2. Each modified sample was loaded into one channel of a 50-channel Helicos Flow Cell, and analyzed using the HeliScope Single Molecule Sequencer. Single molecule reads were mapped to the human genome and binding events were detected by comparing read counts between the two samples in every location of the human genome. Results from a representative section of Chromosome 11 are shown; blue line: immunoprecipitated  DNA from stem cell sample; red line: input control DNA sample. Protein binding sites are clearly visible as sharp peaks in the stem cell sample.

The Approach
Immunoprecipitated genomic DNA from a ChIP experiment is modified with a poly-A tail and loaded onto the instrument. No ligation or PCR amplification steps are required. The tailed fragments hybridize to complementary poly-T strands anchored to the flow cell surface. Inside the HeliScope™ Single Molecule Sequencer, a series of nucleotide addition and detection cycles determine the sequence of each fragment. Open source data analysis software aligns the hundreds of millions of reads to a reference sequence.

Figure 2. Helicos DNA Sample Preparation Methodology for ChIP-Seq

Helicos ChIP-Seq – Providing you with a virtually unbiased, genome-wide view of protein/DNA interactions.