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Prokaryotic RNA Sequencing

Introduction to Prokaryotic RNA Sequencing

Prokaryotic RNA sequencing hyphenates next-generation high-throughput sequencing (NGS) technology to sequence the prokaryotic transcriptome and quickly and systematically procure data of all transcripts (coding and non-coding mRNA).

Prokaryotic RNA sequencing profiles the transcriptome through a stranded RNA library, delivering a more accurate estimation of transcript expression.

Application of Prokaryotic RNA Sequencing

 

  • For profiling transcripts in prokaryotes
  • For predicting operon, TSS, promoter, and UTR
  • For exploring RNA regulation mechanism through antisense transcripts prediction and sRNA analysis

Benefits of Novogene Prokaryotic RNA Sequencing

 

  • Novogene adopts rRNA depletion method for prokaryotic RNA library construction. Effective depletion methods are adopted for a variety of species.
  • Beside gene expression quantification, multiple gene structure and gene function analysis are included in the standard analysis. Customized analysis is also applicable for prokaryotic RNA sequencing.

Prokaryotic RNA-seq Specifications: RNA Sample Requirements

Library Type Sample Type Amount RNA Integrity Number
(Agilent 2100)
Purity
(NanoDrop)
Prokaryotic RNA Library Total RNA ≥ 2μg ≥ 6.0, smooth base line OD260/280 ≥ 2.0;

OD260/230 ≥ 2.0

Prokaryotic RNA -seq Specifications: Sequencing & Analysis

Sequencing Platform Illumina NovaSeq 6000 Sequencing System
Read Length Paired-end 150bp
Recommended Data Amount ≥ 2Gb raw data per sample for the species with reference genome
Content of Data Analysis
  • Data Quality Control
  • Alignment to Reference Genome
  • Gene Expression Analysis
  • Novel Transcript Prediction
  • Gene expression quantification & Differential expression analysis
  • Enrichment Analysis
  • Operon, Promoter and TSS/TTS Prediction
  • SD Sequence Prediction
  • Rho-independent Terminator Sequence Prediction
  • sRNA Secondary Structure Prediction
  • sRNA Targeted Gene Prediction

Novogene Workflow of Prokaryotic RNA -seq 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, an appropriate library is constructed 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 goes through 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 elaborates the step-by-step protocol.

 

[1]https://www.nature.com/articles/jhg2013114

Distributions of gene expression levels

To compare gene expression levels under different conditions, gene expression level and FPKM distribution among different samples are displayed.

Note:
The figure reveals the FPKM density distribution. The x-axis shows the log10(FPKM+1) and the y-axis shows describes the gene density


Volcano plot for differentially expressed genes

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

 

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


UTR length distributions

The 5′ and 3′ UTR sequences were are extracted based on the start and end positions of both transcription and translation. The length distributions of both 5′ and 3′ UTR sequences were are plotted, respectively.

Note:
The x-axis shows the length intervals of UTRs, and the y y-axis shows densities of UTRs in different length intervals. The dotted red dashed line shows describes the averaged length.

*Please contact us to get the full demo report.