High Pressure Sample Preparation Instrumentation for DNA Sequencing

DESCRIPTION provided by applicant Developments in next generation sequencing NGS continue to increase throughput and reduce the costs for whole genome sequencing Unfortunately DNA preparation methods lag behind in both throughput and DNA quality slowing the pace of adoption of new sequencing techniques This Phase II proposal focuses on the development of an automated commercial prototype DNA shearing device that matches the requirements of many popular and emerging NGS platforms In Phase II we will develop a miniature self cleaning active shearing nozzle and integrate it with a constant pressure pump to allow for a flow through shearing design and easy sample introduction and collection In addition we will build on our progress from Phase I to optimize the range of parameters pressure flow rate orifice diameter and design necessary for optimal DNA shearing to generate fragments in predictable narrow size ranges We will manufacture two prototype instruments for placement at our Randamp D laboratory and at a collaborator site and will characterize the performance of the automated system prototypes We have identified vendors for critical system components and secured academic collaborators to validate the performance of the new platform We expect to achieve manufacturing readiness by the conclusion of this Phase II period In Specific Aim we will develop a miniature automatic self cleaning active shearing nozzle followed by Specific Aim where we will integrate the shearing nozzle with a constant pressure pump and the means of sample introduction and collection In this Aim we will also develop computer software to facilitate automatic control of the active nozzle and sample management In Specific Aim we will identify the range of parameters pressure flow temperature and viscosity necessary for optimal performance of proposed integrated system In Specific Aim we will characterize the performance of the automated system prototypes for DNA shearing applications and develop corresponding applications Finally in Specific Aim we will evaluate the suitability of the system for cell lysis and homogenization and optimize the hardware components for that purpose We have identified vendors for most critical components and secured academic collaborators to validate performance of the new platform therefore we expect to achieve the manufacturing readiness by the conclusion of this Phase II periodPUBLIC HEALTH RELEVANCE The aim of this Phase II proposal is to develop a high throughput high pressure automatable hydrodynamic DNA shearing device prototype This device is designed to improve control of DNA fragmentation and minimize losses and potential chemical DNA damage while reducing costs for Next Generation Sequencing and other applications More efficient generation of DNA in the desired size range s will improve DNA sequencing based personalized diagnostics and lead to considerable benefits in biomedical research and healthcare