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Metatranscriptome Sequencing

Introduction to Metatranscriptome Sequencing

Metatranscriptome refers to the study of the function of an entire set of transcripts by RNA-sequencing from environmental samples at a specific time. It tells us about the genes that are highly expressed in a particular microbial environment.

Metatranscriptome sequencing delivers the whole expression profile by randomly sequencing mRNAs and tracks the dynamics of gene expression patterns in microbial communities over time using next-generation sequencing (NGS). This improves our understanding of the structure, function, and adaptive mechanisms of complex communities.

Applications of Metatranscriptome Sequencing

 

  • Functional active bacteria characterization and community metabolic interaction in microbial ecology research
  • Contribution to clinical research such as recognizing immune response recognition and triggering autoimmune processes and immune recognition and autoimmune processes trigger
  • Discovery on innovative resources for drug delivery that are not easily accessible owing to the technical limitations in microbial isolation
  • Improving the quality of food, and provide data related to the metabolic activities of microbial communities.

Benefits of Metatranscriptome Sequencing

 

  • In profiling transcriptome of a natural community, without the isolation and cultivation of microbes
  • In identifying novel genes and detecting the changes in complex microbial communities at the transcriptional level

Metatranscriptome Sequencing Specifications: RNA Sample Requirements

Library Type Sample Type Amount RNA Integrity Number
(Agilent 2100)
Purity
(NanoDrop)
Meta-transcriptome Library Total RNA ≥ 2μg ≥ 6.5, smooth base line OD260/280 ≥ 2.0;

OD260/230 ≥ 2.0;

Metatranscriptome Sequencing Specifications: Sequencing & Analysis

Sequencing Platform Illumina NovaSeq 6000 Sequencing System
Read length Paired-end 150bp
Recommended Data Amount ≥ 20 million read pairs per sample
Content of Data Analysis

Novogene Workflow of Metatranscriptome Sequencing Service

The first step of the project workflow includes sample quality control (Sample QC) to ensure that your samples meet the criteria of the RNA-Seq technique. Then, the appropriate library is prepared according to your target organism and subsequently tested for its quality (Library QC). Next, a paired-end 150 bp sequencing strategy is used to sequence the samples and the resulting data go quality data control (Data QC) to guarantee the quality of the resulting data. Finally, bioinformatic analyses are performed and publication-ready results are provided. The following flowsheet describes the step-by-step protocol.

Featured Publications of Metatranscriptome Sequencing

Length distribution of Unigenes

Unigene: the longest transcript of each cluster from Metatranscriptome assembly.

Note:
The figure shows the Unigene length distribution, number of Unigene in different length intervals.


Species abundance clustering

From each sample, 35 dominant genera were selected based on the species annotation results and abundance information followed by clustering depending on the taxonomy information and the inter-sample differences among samples.

 

Note:
In this figure, the Bray-Curtis distance clustering tree structure is on the left side; the relative abundance distribution map of each sample at the phylum level is shown on the right side.


Comparison of gene expression levels under experimental conditions

The density distribution map and box plot were used to examine the distribution of genes expression in FPKM under different experimental conditions.

 

Note:
In the FPKM box plot, the x-axis shows the sample names, the y-axis is log10 (FPKM+1), and the box plot for each region has five statistics (from top to bottom are the maximum, the upper quartile, median, lower quartile, and minimum). The graph measures total expression and the differences between samples in terms of discreteness.


Volcano diagram of differentially expressed genes

Volcano plots are used to infer the overall distribution of differentially expressed genes.

Note:
Red, and green show significantly up-, and down-regulated genes, respectively. Blue indicates no significance.

 

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