Metatranscriptomic Seq

Metatranscriptomic Seq, or metatranscriptomic sequencing, enables the study of gene expression in complex microbial communities without the need to culture them first. The analysis covers RNA from samples including soil, water, faeces, intestinal contents and swabs.

The resulting data can be used to characterise the community's transcriptional profile, assign biological functions to transcripts and compare gene expression between experimental conditions. The method is used to study microbiome activity and its response to environmental changes or host influences.

Metatranscriptomic Seq workflow

01Sample collection and preservation: Material is collected from the environment under study and preserved to limit RNA degradation and changes in the expression profile after collection.

02RNA isolation and quality control: Total RNA is isolated from the sample. Quality control includes assessing its amount, purity and integrity.

03Library preparation: RNA undergoes rRNA depletion, and is then reverse transcribed into cDNA. Libraries are prepared with adapters and indexes that allow multiple samples to be sequenced together.

04Sequencing: Libraries are sequenced on the Illumina platform using 150 bp paired-end sequencing, meaning that both ends of each fragment are read.

05Bioinformatics analysis: The data undergo quality control, transcript assembly, taxonomic and functional annotation and gene expression analysis.

Applications of Metatranscriptomic Seq

01Human and animal microbiome studies: Analysis of gene expression in microorganisms inhabiting sites such as the gut, oral cavity or skin, and of changes associated with diet or other experimental factors.

02Environmental studies: Assessment of the transcriptional response of microbial communities in soil, water and other environments, for example to changes in conditions or the presence of pollutants.

03Agriculture and plant-microorganism interactions: Studying gene expression in plant-associated microorganisms, including rhizosphere communities, under different growth and stress conditions.

04Fermentation and biotechnology: Monitoring changes in gene expression in communities involved in fermentation processes and analysing genes associated with substrate utilisation and enzyme production.

Technical specifications of Metatranscriptomic Seq

Platform: Illumina.
Sequencing mode: PE150, i.e. 2 × 150 bp.
Library preparation: stranded libraries following rRNA depletion.
Recommended amount of data: 12 Gb per sample.

12 Gb corresponds to approximately 40 million PE150 read pairs with full-length reads. We select the target data output based on microbiome complexity, the proportion of host RNA and the aim of the study. The number of reads usable for analysis also depends on the effectiveness of rRNA removal and data filtering.

Sample Requirements

SAMPLE REQUIREMENTS - DNA/RNA7 items

Parameter	Requirement
Sample type	Total RNA
Total amount	≥1 µg
Concentration	≥50 ng/µl
Volume	≥20 µl
Integrity	RIN ≥6.5
Purity (A260/A280)	1,8-2,0
Purity (A260/A230)	1,0-2,5

RNA isolation from the supplied material is also available. The type, amount and preservation method of samples for RNA isolation must be agreed in advance. Quality control determines whether the material is suitable for library preparation.