ShEEP Request for Single Cell System

Va New Jersey Health Care System, East Orange NJ

Investigators

Abstract

PROJECT ABSTRACT This Shared Equipment Evaluation Program (ShEEP) proposal requests a single-cell library preparation system as a common resource for VA New Jersey Healthcare System (VANJHCS) Investigators. Comprising (1) Namocell Pala Single Cell Dispenser and (2) 10x Genomics Chromium Connect, this assembled single cell system will provide five major VA-Merit Award researchers and three minor VANJHCS researchers the ability to efficiently and reproducibly prepare single cell RNA libraries for sequencing (scRNA-seq), across a diverse group of VA research priorities in: traumatic brain injury (Drs. Yun-Beom Choi, Bruce Citron, and Vedad Delic), neurodegeneration & immunology in Parkinson's disease (Drs. James Park and Wei Lu), and melanoma treated with glutaminergic & checkpoint inhibition (Dr. Suzie Chen), bile acid signaling in infiltrating immune cells during hepatocellular carcinoma development and progression (Dr. Grace Guo), and T-cell biology such as fate differentiation during infection (Dr. Hai-Hui (Howard) Xue). The capabilities of this single-cell RNA library preparation system are not available within VANJHCS and are critical to ongoing and planned research projects. Human disease begins with the cell. An aberrant transcriptome is one of the first manifestations of incipient disease but is often masked or convolved by cellular heterogeneity and bulk sequencing. Single cell analysis reveals insights that are hidden in bulk cell analysis. scRNA-seq has been widely employed to deconvolve under- appreciated cellular heterogeneity, dissect gene regulatory networks, and reveal novel mechanistic insights necessary to improve diagnostics and therapeutics. Because of these considerations, single cell analysis is positioned at the center of a modern high-resolution biomedical investigational tool kit. Reproducible and efficient single cell library preparation has become necessary. Reproducibility stems from Namocell Pala Single Cell Dispenser's ability to reliably screen for fully dissociated and viable single cells. High quality single cell input from Pala Single Cell Dispenser for 10x Genomics Connect is an essential consideration in nervous system, hepatic, and dermal tissues in which cells are networked by extracellular matrices making single cell preparation challenging and variable. 10x Genomics Chromium Connect's efficiency stems from the robot's ability to save an entire working day of investigator liquid-handling time per single cell preparation run. 10x Connect's liquid- handling consistency ensures the reproducibility of scRNA data. As a bonus, Pala Single Cell Dispenser not only serves upstream in the library preparation workflow by screening out cellular doublets, debris, and death, the Pala Single Cell Dispenser can also perform double-duty downstream of 10x Connect by efficiently selecting for CRISPR-edited single cells, saving a month of investigator-CRISPR screening time. Designing and selecting CRISPR-edited single cells is often a logical follow-up experiment after scRNA-seq analysis and interpretation. In summary, the availability of a single cell RNA library preparation system and the ability to efficiently generate CRISPR-edited single cells will further enable Merit-funded VANJHCS investigators to conduct high-resolution research on their respective diseases and identify pathways for translational applications in diagnosis, prognosis, and therapeutic strategies. These endeavors will substantially contribute to the long-term research goals of the VANJHCS. All VANJHCS Merit projects have significant implications for health, and the single cell RNA library preparation system we are requesting will facilitate the progress of these productive projects, as well as further stimulate an esprit de corps in VANJHCS Merit research programs on diseases of high relevance to Veterans.