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Whole Transcriptome Sequencing

Overview

Novogene’s Whole Transcriptome Sequencing service equips the researcher with cutting-edge NGS solutions that provide in-depth bioinformatic analysis on all transcripts including mRNAs and non-coding RNAs. This competitive approach investigates and explores potential transcriptional and regulatory network mechanisms, while providing key insights into interaction and intersection functionality from a comprehensive transcriptomic perspective.

Service Specifications

Applications

  • Co-localization and co-expression of ncRNA and mRNA detection
  • miRNA sponge and target regulatory elements detection
  • Regulatory network investigation: ceRNA regulatory network set up based on lncRNA/circRNA-miRNA-gene pairs, taking lncRNA/circRNA as decoy, miRNA as core and mRNA as target

Advantages

  • Extensive experience with thousands of samples being successfully sequenced.
  • Unsurpassed data quality with a guaranteed Q30 score ≥ 80% that exceeds Illumina’s official benchmarks.
  • Comprehensive analysis using mainstream software and mature in-house pipeline to meet multiple bioinformatic requests.

Sample Requirements

 

Library Type Sample Type Amount RNA Integrity Number
(Agilent 2100)
Purity
(NanoPore)
lncRNA Library & small RNA Library Total RNA ≥ 5 μg Animal ≥ 7.5, Plant ≥ 7, with smooth baseline OD260/280 = 1.8-2.2;
OD260/230 ≥ 1.8;

Sequencing Parameter and Analysis

Platform Illumina Novaseq 6000
Read length Paired-end 150 & Single-end 50
Recommended Sequencing Depth ≥ 40 million read pair per sample (lncRNA library);
≥ 20 million read pair per sample (small RNA library);
Association Analysis of Transcriptome
  • Interaction of lncRNA and miRNA
  • Interaction of mRNA and miRNA
  • Interaction of circRNA and miRNA
  • Regulatory Network of lncRNA, miRNA and mRNA
  • Regulatory Network of circRNA, miRNA and mRNA
  • Note: For detailed information, please refer to the Service Specifications and contact us for customized requests.

    Project Workflow

    Identification of the Spinal Expression Profile of Non-coding RNAs Involved in Neuropathic Pain Following Spared Nerve Injury by Sequence Analysis

    Background:

    Neuropathic pain (NP) is one type of chronic pain and caused by primary damage and dysfunction of the nervous system, which is characterized by dysesthesia, hyperalgesia, and Allodynia and associated with gene expression changes in the sensory pathway. However, the molecular mechanisms are not fully understood and a better understanding of the genetic and various neurobiological bases could provide the clinician with important diagnosis and treatment tools.

    Sampling:

    Total RNA was extracted from the spinal cord dorsal horn tissue

    Sequencing Strategy:

    1. lncRNA library: NEBNext UltraTM Directional RNA Library Prep Kit for Illumina
    2. small RNA library: NEBNext Multiplex Small RNA Library Prep Set for Illumina
    3. sequenced on Illumina HiSeq 2500 platform, 125 bp paired-end and 50-bp single-end reads, respectively.

    Figure 1. The expression profiling changes of lncRNAs in spinal cord of SNI rats Vocalno Plot indicate up and down regulated lncRNAs of rats in group SNI compared with group CON (A)

    Figure 2. The expression profiling changes of circRNAs in spinal cord of SNI rats Vocalno Plot indicate up and down regulated circRNAs of rats in group SNI compared with group CON (A)

    Figure 3. The expression profiling changes of miRNAs in spinal cord of SNI rats Vocalno Plot indicate up and down regulated miRNAs of rats in group SNI compared with group CON (A)

    Figure 4. The expression profiling changes of mRNAs in spinal cord of SNI rats Vocalno Plot indicate up and down regulated mRNAs of rats in group SNI compared with group CON (A)

    Figure 5. Counts of relatived ncRNAs and mRNAs in spinal cord of SNI rats.

    Figure 6. lncRNA-micRNA-mRNAs regulatory network analysis of ncRNAs in spinal cord of SNI rats.

    Figure 7. cirRNA-micRNA-mRNAs regulatory network analysis of ncRNAs in spinal cord of SNI rats.
    Conclusion:

    This study comprehensively identifies regulated ncRNAs of the spinal cord and to demonstrate the involvement of different ncRNA expression patterns in the spinal cord of NP pathogenesis by sequence analysis. This information will enable further research on the pathogenesis of NP and facilitate the development of novel NP therapeutics targeting ncRNAs.


    mRNA-miRNA


    lncRNA-mRNA


    circRNA-mRNA


    mRNA-miRNA-lncRNA