Award Date

August 2025

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Life Sciences

First Committee Member

Martin R. Schiller

Second Committee Member

Brian Hedlund

Third Committee Member

Mira Han

Fourth Committee Member

Katherine Shortt

Fifth Committee Member

Sarah Harris

Number of Pages

399

Abstract

Understanding the functional consequences of genetic mutations remains a central challenge in modern biology, with far-reaching implications for human health and disease. While early systematic methods like alanine scanning and phage display provided foundational insights into protein structure and function, the emergence of high-throughput approaches—such as Multiplexed Assays of Variant Effect (MAVEs)—and predictive tools powered by artificial intelligence have vastly expanded our ability to profile mutational landscapes. However, these methods are often constrained by trade-offs between accuracy, scalability, and biological relevance.This dissertation presents the development and application of the GigaAssay, the world’s first high-throughput functional assay capable of delivering both high accuracy and scalability. Built upon a modular, one-pot experimental framework, the GigaAssay enables the quantitative assessment of thousands of mutations simultaneously, while maintaining single-molecule resolution through the use of hundreds of unique molecular identifier (UMI) barcodes per variant. The technology's generalizability and robustness are demonstrated by exploring the mutation space of two vastly different proteins, HIV-1 Tat and HER2. In addition to detailing the experimental and computational innovations that underpin the GigaAssay, this work highlights its transformative applications in virology and oncology, offering new avenues for functional genomics, drug development, and precision medicine. By enabling systematic and reproducible functional interrogation of genetic variation at unprecedented scale and accuracy, the GigaAssay empowers a new era of biological discovery.

Keywords

Function; GigaAssay; HER2; HIV; Mutagenesis; Protein

Disciplines

Biochemistry | Bioinformatics | Biostatistics

File Format

pdf

Degree Grantor

University of Nevada, Las Vegas

Language

English

Rights

IN COPYRIGHT. For more information about this rights statement, please visit http://rightsstatements.org/vocab/InC/1.0/

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