Ultralow Input RNA-Seq

CD Genomics is proud to provide ultralow input RNA-seq service using leading-edge Illumina sequencing techniques. The highly sensitive ultralow input RNA-Seq service enables researchers to explore the gene expression profile with the ultra-low amount of input RNA. This technique offers a powerful tool for analyzing the gene expression in cancer research and studies with extremely rare samples. CD Genomics is committed to delivering the highest quality, even with difficult templates, and providing prompt customer assistance at all times.

Overview

Samples that contain distinct cell types exhibit much variation, therefore, selecting cell clusters that you want to study for RNA sequencing becomes more informative and easier to interpret. Ultralow input RNA sequencing (RNA-seq) is a powerful tool for gene expression profiling with limited number of cells, especially when cell-to-cell variation are not the aim. It allows you to detect splice junctions and significant changes in gene expression for low abundant transcripts, as well as discovery of biomarkers and transcription-factors. CD Genomics offers a simple, low-cost, and low-bias ultralow input RNA-seq with Illumina sequencing. Combining the experimental and computational optimization, our solution can be comparable with the bulk RNA-seq, minimizing the bias with much less mappable reads. Our service is specifically designed for input amounts down to 2 pg, applicable for good quality RNA of challenging samples. Our expert team ensures you uniform transcript coverage even with ultralow input amount. Built on a foundation of unmatched expertise in transcriptomics, CD Genomics is highly experienced in ultralow input RNA-seq and bioinformatic analysis.

Features

UMIs Ultralow Input Competitive Pricing Bioinformatics
Unique Molecular Identifiers (UMIs) are ligated to the RNA molecules to minimize PCR duplication rate. Minimally validated RNA input: 2 pg; Ideally requiring more RNA for QC and normalization. Provide raw data as well as summary data with turn-around time and cost. Complete bioinformatics support, customized for your need.

Project Workflow

Sample Preparation

1. Sample Preparation

RNA purification;
quality assessment and quantification

Library Preparation

2. Library Preparation

cDNA stranded library preparation;
library QC.

Sequencing

3. Sequencing

llumina NovaSeq 6000;
PE 50/75/100/150;
Quality score Q30 of ≥80%,
generally 20 million reads.

Data Analysis

4. Data Analysis

Visualize and preprocess results, and perform custom bioinformatics analysis.

Bioinformatics Analysis Pipeline

Ultralow Input RNA-Seq

In-depth data analysis:

  • Biostatistical analysis –transcript copy number comparisons, expression levels (FPKM values) per gene, length distribution, multi-parameter data analysis, etc.
  • Differential gene-expression analysis
  • RNA structure analysis
  • Target gene prediction and functional analysis
  • Detecting novel and rare transcripts and mutations
  • Novel transcript prediction
  • PCA plot, heatmap, pathway analysis

Sample Requirements

RNA sample (quantity ≥ 2 pg, ideally deliver as much RNA as possible for QC and normalization)

1.8 ≤ OD260/280 ≤ 2.2, OD260/230 ≥ 2.0, RIN ≥ 7.0, 28S:18S ≥ 1.0.

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. Palomares M A, Dalmasso C, Bonnet E, et al. Systematic analysis of TruSeq, SMARTer and SMARTer Ultra-Low RNA-seq kits for standard, low and ultra-low quantity samples. Scientific reports, 2019, 9(1): 1-12.
  2. Curion F, Handel A E, Attar M, et al. Targeted RNA sequencing enhances gene expression profiling of ultra-low input samples. RNA biology, 2020: 1-13.
* For Research Use Only. Not for use in diagnostic procedures.


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