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High-Throughput Transcriptome Analysis for Investigating Host-Pathogen Interactions
Published on: March 5, 2022
Enriching pathogen transcripts from infected samples: a capture-based approach to enhanced host-pathogen RNA
Zachary W Bent1, Mary B Tran-Gyamfi, Stanley A Langevin
1Sandia National Laboratories, Livermore, CA 94551, USA.
Analytical Biochemistry
|March 29, 2013
Summary
A new RNA capture technique significantly enriches pathogen transcripts from infected samples. This method improves pathogen gene analysis by increasing sequencing depth and coverage, crucial for understanding host-pathogen interactions.
Area of Science:
- Microbiology
- Genomics
- Infectious Diseases
Background:
- Understanding host-pathogen interactions requires analyzing RNA transcripts from both organisms during infection.
- Analyzing pathogen RNA is challenging due to the high abundance of host RNA in infected samples.
- Second-generation sequencing often yields insufficient pathogen reads for effective gene-level analysis.
Purpose of the Study:
- To develop and validate a novel capture-based technique for enriching pathogen transcripts.
- To overcome the limitations of host RNA dominance in transcriptomic studies of infection.
- To enable more comprehensive gene-level analysis of pathogens in infected hosts.
Main Methods:
- A novel capture-based technique and device were developed to enrich pathogen transcripts.
- Time-course tissue culture infections were performed using Rift Valley fever virus and Francisella tularensis.
- RNA sequencing (RNA-Seq) was conducted on treated and untreated samples for comparative analysis.
Main Results:
- The capture technique enriched pathogen reads by over an order of magnitude in all tested cases.
- Significant increases were observed in the number of pathogen genes detected, their coverage, and sequencing depth.
- The method demonstrated efficacy in enriching transcripts for both viral and bacterial pathogens.
Conclusions:
- The novel capture technique substantially enhances the proportion of pathogen RNA in sequencing libraries.
- This method significantly improves the ability to perform gene-level analysis of pathogens during infection.
- The versatile technique holds promise for studying a wide range of host-pathogen systems.
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