Dual RNA Sequencing

At CD Genomics we've been providing dual RNA sequencing services for over 10 years to support researchers, institutes, and biotechnology and pharmaceutical companies all over the world with diverse needs. Offering a total transcriptomic solution based on our extensive experiences and know-how accumulated over the years, CD Genomics leads the field of RNA sequencing utilizing NGS, Oxford nanopore, and Pacific Biosciences technologies.

Overview

Eukaryotic host cells are subject to infection by various agents, from viruses to bacteria to parasites like fungi and protozoa. Host-pathogen interaction initiates gene expression changes within both interacting organisms, leading to the adaptation and persistence, or clearance of the pathogen. The determination of the associated gene expression changes in both the host cell and the pathogen provides insights into a comprehensive understanding of host-pathogen interactions, including identification of novel virulence factors in the pathogens and novel pathway in responses to pathogen exposure in the host. Dual RNA sequencing (RNA-seq) is emerging as a powerful tool for the dissection of the in vivo host-pathogen interplay, uncovering the influences that both organisms exert over each other. Dual RNA-seq is an ideal tool for elucidating the molecular dynamics underlying bacterial fitness and host cell phenotype among multiple divergent host cell lineages. It allows to simultaneously detect host- and pathogen-specific transcripts in mixed samples and detect low-abundance transcripts. This newfound feasibility has enabled the direct association of pathogenic gene activity with a specific host response, as well as the identification of pathogenic phenotypes that are invisible in standard virulence assays. This global, sensitive, and unbiased method has been successfully applied on several plant-pathogen models, exhibiting suitability and superiority compared with traditionally used techniques.

Features

Extensive Experiences Stranded Library Competitive Pricing Bioinformatics
We have been providing RNA-seq services since 2014 and have accumulated much experience. Strand information resolves read ambiguity in overlapping genes transcribed from opposite strands. Provide raw data as well as summary data with turn-around time and cost. Complete bioinformatics support, customized for your need.

Project Workflow

Sample Preparation

1. Sample Preparation

RNA purification;
quality assessment and quantification

Library Preparation

2. Library Preparation

cDNA stranded library preparation;
library QC

Sequencing

3. Sequencing

llumina NovaSeq 6000;
PE 50/75/100/150;
Quality score Q30 of ≥80%

Data Analysis

4. Data Analysis

Visualize and preprocess results, and perform custom bioinformatics analysis

Bioinformatics Analysis Pipeline

Dual RNA Sequencing

In-depth data analysis:

  • Parallel read mapping against the respective host and pathogen genomes
  • Quantification and normalization
  • Differential expression analysis
  • Identification of molecular phenotypes of pathogenic genes
  • PCA plot, heatmap, pathway analysis, network inference

Sample Requirements

RNA sample (quantity ≥ 2 μg)

1.8 ≤ OD260/280 ≤ 2.2, OD260/230 ≥ 2.0, RIN ≥ 7.0, 28S:18S ≥ 1.0.

Sample storage: RNA can be dissolved in ethanol or RNA-free ultra-pure water and stored at -80°C. RNA should avoid repeated freezing and thawing.

Shipping Method: When shipping RNA samples, the RNA sample is stored in a 1.5 mL Eppendorf tube, sealed with sealing film. Shipments are generally recommended to contain 5-10 pounds of dry ice per 24 hours.

Deliverable: FastQ, BAM, coverage summary, QC report, custom bioinformatics analysis.

References:

  1. Westermann A J, Gorski S A, Vogel J. Dual RNA-seq of pathogen and host. Nature Reviews Microbiology, 2012, 10(9): 618-630.
  2. Westermann A J, Förstner K U, Amman F, et al. Dual RNA-seq unveils noncoding RNA functions in host–pathogen interactions. Nature, 2016, 529(7587): 496-501.
* For Research Use Only. Not for use in diagnostic procedures.


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