Exosomal Long Non-coding RNA (lncRNA) Sequencing Service

Q: Q1: How does exosomal lncRNA sequencing distinguish between functional lncRNAs and non-functional RNA fragments?

A: We use a multi-criteria filtering approach: (1) Length verification (retaining transcripts >200 nt, excluding shorter fragments). (2) Coding potential analysis (CPAT/CPC2 tools to confirm non-protein-coding nature). (3) Expression consistency (requiring detection in ≥3 biological replicates). (4) Conservation analysis (prioritizing lncRNAs with evolutionary conservation across species). This ensures we focus on functionally relevant lncRNAs, unlike small RNA sequencing (which often includes non-functional RNA degradation products).

Q: Q2: Can your service identify lncRNA splice variants, and why is this important for exosome research?

A: Yes. Our long-fragment library preparation protocol (300-500 bp) and high sequencing read depth (40-60 million reads) enable effective detection of lncRNA splicing variants, which is crucial. Exosomes often encapsulate specific splicing variants with unique regulatory functions. For instance, in cancer exosomes, a truncated variant of the lncRNA MALAT1 may lack the miRNA-binding domain, thereby altering its ability to regulate target genes. mRNA sequencing primarily focuses on splicing variants encoding proteins, while small RNA sequencing cannot capture long transcripts. This highlights the unique advantage of our lncRNA sequencing service.

Q: Q3: How do you quantify exosomal lncRNA expression, and is it comparable to mRNA quantification?

A: We use TPM (Transcripts Per Million) for lncRNA quantification, a normalized metric that accounts for transcript length and sequencing depth. This allows cross-sample comparison. Unlike mRNA (which has consistent polyA tails for reliable quantification), lncRNAs vary in polyadenylation, so we avoid methods like RPKM (which biases against non-polyadenylated transcripts). Our approach ensures accurate lncRNA quantification, with results directly comparable to mRNA data if integrating multi-omics datasets.

Q: Q4: Can exosomal lncRNA sequencing reveal tissue-of-origin for exosomes, and how?

Yes. Many long non-coding RNAs (lncRNAs) exhibit tissue specificity; for example, lncRNA H19 is enriched in liver-derived exosomes, while lncRNA NEAT1 is more abundant in brain-derived exosomes. We predict the tissue origin of exosomes by comparing lncRNA expression profiles against our proprietary tissue-specific lncRNA database. This approach holds significant implications for disease research (e.g., identifying the primary tumor source of circulating cancer exosomes) and offers greater specificity than mRNA-based tissue tracing techniques, which typically rely on widely expressed reference genes.

Exosomal long non-coding RNAs (lncRNAs) are >200 nt non-protein-coding RNAs enriched in exosomes, acting as key regulators of gene expression, cell signaling, and intercellular communication. Creative Biostructure's exosome-based lncRNA sequencing service captures full-length lncRNAs, revealing their roles in disease progression, tissue repair, and therapeutic response. Leveraging IncRNA-specific sequencing workflows and advanced bioinformatics tools, we deliver high-precision data and comprehensive lncRNA annotation.

What is Exosomal lncRNA Sequencing and Why is it important?

Exosomal lncRNA sequencing is a next-generation sequencing (NGS) technique used to comprehensively detect and profile long non-coding RNAs (lncRNAs) found in exosomes, which are small extracellular vesicles that facilitate intercellular communication. By quantifying the expression levels of these exosomal lncRNAs, researchers can elucidate their functional roles in various physiological and pathological processes, especially in cancer. As a result, they represent highly promising biomarkers for both diagnostic and prognostic applications.

Exosomal lncRNA signature. Figure. 1 An exosome-related lncRNA signature. (Peng W, et al. 2023)

Our Exosomal lncRNA Sequencing Services

Long non-coding RNAs (lncRNAs) are key players in gene regulatory networks. However, their low abundance, complex splice variants, and lack of a polyadenylated (polyA) tail make their capture and characterization challenging. Creative Biostructure offers professional end-to-end lncRNA sequencing solutions to help customers overcome these challenges. From experimental design, sample quality control, and sequencing operations to data analysis, our expert team provides comprehensive technical support throughout the entire process to ensure high-quality results for every project.

We employ the Illumina NovaSeq 6000 platform combined with an lncRNA-optimized protocol to provide the following technologies and services tailored to the characteristics of long non-coding RNAs:

Technique	Description
rRNA Depletion Technology	This method ensures≥95% rRNA removal and comprehensive lncRNA capture by using probe-based rRNA removal (targeting 18S/28S rRNA) rather than polyA+ enrichment, which misses most lncRNAs due to their lack of polyA tails.
Long-fragment Library Construction	Tailored to the structural characteristics of lncRNAs, we have optimized the library construction process to preferentially obtain long fragments of 300-500 bp. This approach maximizes the preservation of complete lncRNA structural information, laying a solid foundation for the accurate analysis of their splice variants and secondary structures.
High Read Depth	40-60 million clean reads per sample, covering low-abundance exosomal lncRNAs (down to 0.5 copies per cell) and enabling accurate detection of lncRNA expression and co-expression patterns.

lncRNA Sequencing Analysis Service Workflow

1

Exosome Isolation (Optional for Raw Samples)

For raw samples (e.g., serum, cell supernatants), use "ultracentrifugation + density gradient centrifugation" to remove lipoprotein and protein contaminants (lncRNA is more sensitive to protein binding than mRNA/small RNA).

2

lncRNA-Specific RNA Extraction

Use RNase-free kits to extract total RNA from exosomes; avoid polyA+ selection (to retain non-polyadenylated lncRNAs); add DNase I to eliminate genomic DNA.
- rRNA Depletion & Library Construction
- Deplete rRNA using species-specific probes.
- Fragment total RNA into 300-500 bp segments (optimized for lncRNA length).
- Synthesize cDNA and add Illumina adapters to construct strand-specific libraries (preserves lncRNA's transcriptional direction, critical for regulatory analysis).

3

High-Throughput Sequencing

Load libraries onto Illumina NovaSeq 6000 for paired-end sequencing (2×150 bp), ensuring sufficient coverage for lncRNA splice variant detection.

4

lncRNA-Focused Data Analysis

- Basic Analysis: FastQC (quality control), STAR (genome alignment with lncRNA annotation), HTSeq (lncRNA expression quantification);
- Advanced Analysis: DESeq2 (differential lncRNA screening), lncRNA annotation (using NONCODE/lncRNAdb), target gene prediction (cis/trans regulation), lncRNA-mRNA/protein interaction network construction, GO/KEGG enrichment of target genes.

Comprehensive lncRNA sequencing and analysis workflow. Figure 2. Workflow of lncRNA sequencing analysis. (Creative Biostructure)

Supported Sample Range

Familiarize yourself with the supported sample types and submission guidelines to ensure your specimens comply with laboratory standards.

Samples	Submit Request
Serum/plasma	≥300 μL, no hemolysis (hemoglobin degrades lncRNA)
Cell culture supernatants	≥15 mL, freshly collected (avoid RNA degradation)
Cerebrospinal fluid	≥150 μL, stored at -80°C immediately
Urine	≥800 μL, add RNA protection agent at collection.

Deliverables

Clients receive a comprehensive, publication-ready report including:

QC Report: Total RNA integrity (RIN value), rRNA depletion efficiency (>95%), library concentration/uniformity, sequencing Q30 score (>95%), alignment rate (>92%).
Raw & Processed Data: FASTQ files (raw reads), BAM files (aligned reads), lncRNA expression matrix (TPM values), lncRNA annotation table (name, location, length, source database).
Advanced Analysis Results:
Differential lncRNA list (fold change ≥2, p<0.05) with volcano plots and heatmaps.
Target gene prediction results (cis/trans targets, binding sites).
lncRNA-mRNA/protein interaction network diagrams (e.g., lncRNA HOTAIR targeting PTEN mRNA).
GO/KEGG enrichment of target genes (revealing lncRNA's regulatory pathways).

Case Study

Case: Improved Exosome Yield from MSCs Using One-Step Sucrose Cushion Ultracentrifugation

Background

The role of exosomal long non-coding RNAs (lncRNAs) in type 1 diabetes (T1DM) remains unclear. This study profiles plasma exosomal lncRNA expression in T1DM to explore their potential functions in disease pathogenesis.

Methods

Exosomal lncRNA expression profiles were detected by Illumina Hiseq platform.
Six exosomal lncRNAs were selected to validate their expression level by using quantitative real-time PCR (qRT-PCR).
Bioinformatics analysis approaches were carried out to explore the potential biological function of differentially expressed lncRNAs.

Conclusion

A total of 162 differentially expressed exosomal lncRNAs were identified in T1DM patients compared with control subjects, among which 77 up-regulated and 85 down-regulated. These results highlighted the potential role of exosomal lncRNAs in T1DM pathogenesis. A better understanding of exosomal lncRNA profiling will provide novel insights into its molecular mechanisms.

Plasma exosomal lncRNA profiling in case and control groups. Figure 3. Plasma-derived exosomal lncRNA profiles of case and control group. (Pang H, et al., 2022)

Creative Biostructure's exosomal lncRNA sequencing service leverages precise strand-specific library preparation and professional bioinformatics workflows. Our integrated solution provides end-to-end support, spanning complete exosome isolation, high-quality library construction, and in-depth data mining. Contact us today to obtain a customized solution that will accelerate your research progress.

References

Peng W, Bai S, Zheng M, et al. An exosome-related lncRNA signature correlates with prognosis, immune microenvironment, and therapeutic responses in hepatocellular carcinoma. Translational Oncology. 2023, 31: 101651.
Pang H, Fan W, Shi X, et al. Characterization of lncRNA profiles of plasma-derived exosomes from type 1 diabetes mellitus. Frontiers in Endocrinology. 2022, 13: 822221.

Frequently Asked Questions

For any inquiries, our support team is ready to help you get technical support for your research and maximize your experience with Creative Biostructure.

Related Services

Related Resources

What is Exosomal RNA Sequencing?

Online Inquiry

Our Valued Partnerships