Cavitation Enhancement of Biospecimen processing for Improved DNA Fragmentation

描述（由申请方提供）：随机、无偏倚的DNA片段化是下一代测序（NGS）和染色质免疫沉淀（ChIP）所必需的。由于DNA片段化对于NGS和ChIP来说都是一个非常有问题的步骤，因此任何提高该步骤效率和一致性的技术对于研究实验室和临床诊断都是非常理想的。此外，一种技术，可以使这一步骤更容易，没有新的设备和非常少的成本，实验室将是理想的。我们建议将脂质包裹的微泡应用于纯化的基因组和甲醛交联样品的DNA片段化。我们最近探索了这项技术的可行性，结果令人印象深刻。初步数据表明，微泡可以大大提高声学DNA片段的一致性。此外，这些气泡以微升体积以每孔1至10美分的成本添加到DNA或细胞悬浮液中，并且可以与任何标准声学超声仪一起使用，与其他技术相比呈现出显著的成本节省以改善DNA剪切。此外，微泡技术大大减少了优化剪切所需的时间，可能大大提高该技术的通量。作为第二个方面，我们还将评估微泡技术的潜力，以提高福尔马林固定石蜡包埋（FFPE），或微活检组织处理。我们的目标将是确定这一现象的最佳性能的条件。检测的变量将包括缓冲试剂、微泡尺寸、微泡浓度、声频、声峰值压力和超声处理持续时间。该项目最后将出版标准作业程序，以传播这一新技术的效用。

DESCRIPTION (provided by applicant): Random, unbiased fragmentation of DNA is necessary for next-generation sequencing (NGS) and chromatin immunoprecipitation (ChIP). Since DNA fragmentation can be a very problematic step for both NGS and ChIP, any technology that increased the efficiency and consistency of this step will be highly desirable for both research laboratories and in clinical diagnostics. Also, a technology that could make this step easier with no new equipment and very little cost to the laboratory would be ideal. We propose to apply the use of lipid encapsulated microbubbles to the fragmentation of DNA from both purified genomic and formaldehyde crosslinked samples. We have recently explored the feasibility of this technology, and our results were impressive. Preliminary data indicate that microbubbles can greatly improve the consistency of acoustic DNA fragmentation. Additionally, these bubbles are added in microliter volumes to the DNA or cell suspension at a cost ranging from one to ten cents per well, and can be used with any standard acoustic sonicator, presenting substantial cost savings compared to other techniques to improve DNA shearing. Furthermore, the microbubble technique greatly reduces the time required to optimize shearing, potentially greatly improving the throughput of this technique. As a second aspect, we will also assess the potential of microbubble technology to enhance tissue processing of formalin-fixed paraffin embedded (FFPE), or microbiopsies. Our goal will be to determine conditions for optimal performance of this phenomenon. Variables tested will include buffer reagents, microbubble size, microbubble concentration, acoustic frequency, acoustic peak pressure, and sonication duration. The project will conclude with publication of Standard Operating Procedures to disseminate the utility of this new technology.