Biofluid microRNA Sequencing

Biofluid microRNA Sequencing is a next-generation sequencing (NGS)-based method to comprehensively detect microRNA extracted from various biofluid samples. It can profile microRNA levels and predict novel microRNAs. Our pioneered biofluid microRNA sequencing ensures the highest level of reproducibility, specificity, sensitivity and accuracy for your project.

Overview

MicroRNAs (miRNAs) are conserved, endogenous, non-coding, small (18–25 nucleotides), single-stranded molecules that regulate gene expression at a post-transcriptional level by binding to the 3' untranslated region (UTR) of mRNA. miRNAs are present in circulating blood plasma, as well as within other biofluids such as exosomes, urine, cerebral spinal fluid and even breast milk. Biofluid miRNAs represent a potential novel approach for diagnostic screening. Indeed, characteristic changes in the miRNA profiles from biofluid have identified unique signatures that could be developed as novel biomarkers for several cancers or other conditions.

We know the challenges in sequencing miRNAs from biofluid samples and overcome this by taking advantage of years of in-house experience in working with liquid biopsies. Over the years, we have conducted thousands of miRNA expression profiling experiments from biofluids such as plasma and exosomes. We combine next-generation sequencing (NGS) and specialized RNA extraction and purification kits for biofluid microRNA sequencing to provide sensitive and accurate miRNA expression profiling, contributing to the discovery and validation of novel non-invasive biomarkers for a range of diseases.

Features

Various Sample Sources Transcriptome-Wide Bioinformatics Analysis One-Stop Solution
This method can be applied to a variety of biofluid samples from humans or other species. Profile all microRNAs, either known or unknown, in your biofluid sample. Our integrated bioinformatics pipeline can be tailored to suit your project. One-stop solution from sample QC and library construction, to sequencing and data analysis.

Project Workflow

Sample Preparation

1. Sample Preparation

qPCR-based QC to detect microRNA content, inhibition and hemolysis

Library Preparation

2. Library Preparation

Size selection (15 – 30 nt); Optimized library preparation protocols for limited RNA content; assessment and quantification.

Sequencing

3. Sequencing

Illumina instruments;
1x 50 bp reads;
7.5 M reads per sample.

Data Analysis

4. Data Analysis

Visualize and preprocess results, and perform custom bioinformatics analysis.

Bioinformatics Analysis Pipeline

Bioinformatics Analysis Pipeline

In-depth data analysis:

  • microRNAs profiling
  • Predict novel microRNAs
  • Detect novel and rare transcript variants
  • Differential expression analysis of microRNAs
  • Target gene prediction and functional analysis of microRNAs
  • GO analysis
  • KEGG analysis
  • Principal Component Analysis (PCA plot) and heatmap

Sample Requirements

Total RNA (concentration ≥ 1 ng/uL, quantity ≥ 20 ng)
1.8 ≤ OD260/280 ≤ 2.2, OD260/230 ≥ 2.0, RIN ≥ 6.5, 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. Alexandrov P N, Dua P, Hill J M, et al. microRNA (miRNA) speciation in Alzheimer’s disease (AD) cerebrospinal fluid (CSF) and extracellular fluid (ECF). Int J Biochem Mol Biol. 2014, 3(4): 365-73.
  2. Gildea J J, Carlson J M, Schoeffel C D, et al. Urinary Exosome miRNome Analysis and its Applications to Salt Sensitivity of Blood Pressure. Clin Biochem. 2013, 46(12): 1131-4.
  3. Mooney C, Raoof R, El-Naggar H, et al. High Throughput qPCR Expression Profiling of Circulating MicroRNAs Reveals Minimal Sexand Sample Timing-Related Variation in Plasma of Healthy Volunteers. PLOS ONE. 2015, 10(12): e0145316.
* For Research Use Only. Not for use in diagnostic procedures.


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