Bacterial Small RNA-Seq

Bacterial small RNA-Seq is a method to comprehensively detect small RNAs based the next-generation sequencing (NGS). It can accurately reveal the complete picture of the transcriptome in a biological sample under specific conditions. We provide bacterial small RNA-seq to help you analyze microRNAs (miRNAs), small interfering RNAs (siRNAs), piwi-interacting RNAs (piRNAs), small nucleolar RNAs (snoRNAs) and small nuclear RNAs (snRNAs) at one time.

Overview

Small RNA is a type of non-protein-coding RNAs that is short in length (<200 nt). Small RNA species generally include miRNA, siRNA, and piRNA, snoRNA, snRNA and so on. Small RNA populations and abundance can vary significantly among different species and tissue types. Generally, small RNAs are formed by fragmentation of longer RNA sequences with the help of biomolecules, such as the dedicated sets of enzymes. Small RNAs perform many biological functions. For example, miRNAs and siRNAs play an important role in post-transcriptional regulation of gene expression. Small RNAs are involved in a number of biological processes including cell proliferation and differentiation, development, and apoptosis.

Bacterial small RNA sequencing (RNA-seq) isolates small RNAs by size fractionation using silica spin columns or gel electrophoresis selection from total RNA, followed by RNA adapter ligation using a 5’ adenylated DNA adapter with a blocked 3’end. Subsequently, small RNAs are reversely transcribed, amplified by PCR and sequenced. Our bacterial small RNA-seq service can obtain a premium small RNA sequencing library, enabling profiling of small RNAs, detection of mutations/variants/isoforms, functional analysis, prediction and characterization of novel small RNAs, and differential expression analysis.

Features

Effective Data Transcriptome-Wide Bioinformatics Analysis One-Stop Solution
Specific enrichment of sRNA (50-300 nt), offering a high proportion of effective data. Profile all miRNAs, tRNAs, snRNAs, snoRNAs, piRNAs, either known or unknown, in your sample. Our integrated bioinformatics pipeline can be tailored to suit your project. One-stop solution from sample QC, library construction, to sequencing and data analysis.

Project Workflow

Sample Preparation

1. Sample Preparation

RNA/cDNA QC;
rRNA removal

Library Preparation

2. Library Preparation

50-300 nt size selection;
strand-specific small RNA cDNA libraries.

Sequencing

3. Sequencing

Illumina HiSeq SE50
>10M reads

Data Analysis

4. Data Analysis

Visualize and preprocess results, and perform custom bioinformatics analysis.

Bioinformatics Analysis Pipeline

Bioinformatics Analysis Pipeline

In-depth data analysis:

  • Small RNA classification and quantification
  • Differential expression analysis of transcripts
  • Target gene prediction and annotation
  • Novel small RNA prediction
  • GO analysis
  • KEGG analysis

Sample Requirements

RNA sample (concentration ≥ 200 ng/uL, quantity ≥ 2 ug)
1.8 ≤ OD260/280 ≤ 2.2, OD260/230 ≥ 2.0, RIN ≥ 7.0, 28S:18S≥1.0.
Please make sure that the RNA is not significantly degraded.

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. Baran-Gale J, Kurtz C L, Erdos M R, et al. Addressing Bias in Small RNA Library Preparation for Sequencing: A New Protocol Recovers MicroRNAs that Evade Capture by Current Methods. Frontiers in Genetics, 2015, 6(e73240).
  2. Buschmann D, Haberberger A, Kirchner B, et al. Toward reliable biomarker signatures in the age of liquid biopsies-how to standardize the small RNA-Seq workflow. Nucleic acids research, 2016, 44(13): 5995-6018.
* For Research Use Only. Not for use in diagnostic procedures.


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