SBIR: Technology for Overcoming Bottlenecks in Chromatin Extraction from Challenging Biological Samples

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Molecular & Cellular Analysis Technologies
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PROJECT SUMMARY The complex organization of chromatin (DNA with its associated nuclear proteins) is closely tied to theregulation of gene expression, and evidence points to a central role for chromatin deregulation in tumordevelopment. A common analysis of chromatin regulatory protein localization is the chromatinimmunoprecipitation (ChIP) assay, which involves crosslinking of proteins to DNA, followed by chromatinfragmentation, and then antibody mediated immunoprecipitation of the DNA fragments. Subsequentquantification (quantitative PCR or Next Generation Sequencing) is usually performed. This assay is a key toolfor both industry and academic research studying cancer-related epigenetics The challenge is that the formaldehyde crosslinks render the chromatin resistant to mechanical (i.e.acoustic sonication) lysis, making current fragmentation techniques inefficient and time-consuming.Furthermore, these techniques result in high sample-to-sample variability and require high-power, low-throughput, and expensive sonication devices. Our customer discovery interviews have highlighted these painpoints; there is an urgent need for a higher quality and higher throughput method of chromatin fragmentationbefore assays such as ChIP can be incorporated into cancer diagnostics applications. Triangle Biotechnology recently developed a unique cavitation enhancing reagent (RapidShear) thatsubstantially improves the efficiency of acoustic fragmentation of genomic DNA. However, this reagentformulation shows little to no improvement for chromatin fragmentation from fixed cells. To address theaforementioned sample preparation challenges in the epigenetics space, we are developing a new reagent,MegaShear, a more aggressive formulation for acoustic cavitation enhancement designed specifically forchromatin processing. Preliminary studies indicated that MegaShear facilitates a substantial improvement inchromatin fragmentation throughput, consistency, and yield. Validation and optimization of this technology willalleviate a critical bottleneck in the ChIP workflow. MegaShear will enable a substantial leap in high qualitysample preparation throughput desired by large companies, as well as an increase in accessibility by enablingthe use of low-cost sonicators, desired by small academic labs. With this SBIR, we will demonstrate proof of concept for MegaShear, validating the ability to producehigh quality chromatin with high consistency, reducing capital equipment cost, and substantially increasingthroughput. Success of this project will enable a novel commercial solution for addressing market painpoints in chromatin analysis.