# Development of Metagenomic RNA-Seq Standard Operating Procedure for Variant-agnostic Whole Genome Sequencing of SARS-CoV-2 isolates from Animal Samples

> **NIH FDA U18** · UNIVERSITY OF MISSOURI-COLUMBIA · 2022 · $97,564

## Abstract

Abstract
The genetic diversity of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in
non-human hosts is more poorly understood. Although whole-genome sequencing (WGS) has
found extensive application in characterizing the genetic diversity of SARS-CoV-2 isolates over
the past two years, current protocols are aimed solely at identifying mutants circulating in human
hosts. This approach could potentially miss new biologically relevant SARS-CoV-2 variants that
may emerge as this virus continuously interacts with disparate non-human hosts. The proposed
research, therefore, focuses on the establishment of a validated standard operating procedure
(SOP) to directly sequence and analyze SARS-CoV-2 in clinical animal specimens using the
metagenomic RNA-Seq approach. Compared with the most widely used multiplex PCR targeted
amplicon sequencing approach (MTA-Seq), this unbiased metagenomic RNA-Seq approach
faithfully covers the entire viral genome and increases the odds of detecting new and unique
variants that may not be present in human populations.
We will use archived SARS-CoV-2 positive samples originated from diagnostic cases to evaluate
two novel library preparation systems, which are known either for the universal reduction of host
background (Zymo-Seq Ribo-Free Total RNA kit) or for enhancement of viral genome
amplification sensitivity with single primer isothermal amplification (SPIA) (Revelo RNA-Seq
Library Kit). The libraries will then be sequenced on both the Illumina iSeq and MiSeq
platforms. We will compare the performance of both kits for WGS on the basis of sequencing
depth, genome coverage, sequencing uniformity, and detection limits. Bioinformatic analyses
will be performed in the public Galaxy server and its GalaxyTrakr instance. The fully validated
SOP developed from this project will be submitted to FDA Vet-LIRN for distribution to other
member laboratories. On completion, this work will significantly enhance the capabilities of Vet-
LIRN laboratories and strengthen collaboration across the network.

## Key facts

- **NIH application ID:** 10610042
- **Project number:** 1U18FD007720-01
- **Recipient organization:** UNIVERSITY OF MISSOURI-COLUMBIA
- **Principal Investigator:** Solomon Olawole Odemuyiwa
- **Activity code:** U18 (R01, R21, SBIR, etc.)
- **Funding institute:** FDA
- **Fiscal year:** 2022
- **Award amount:** $97,564
- **Award type:** 1
- **Project period:** 2022-09-01 → 2024-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10610042

## Citation

> US National Institutes of Health, RePORTER application 10610042, Development of Metagenomic RNA-Seq Standard Operating Procedure for Variant-agnostic Whole Genome Sequencing of SARS-CoV-2 isolates from Animal Samples (1U18FD007720-01). Retrieved via AI Analytics 2026-05-27 from https://api.ai-analytics.org/grant/nih/10610042. Licensed CC0.

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