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Small RNA Sequencing

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To support the increasing research interest in small RNA, CD Genomics is offering small RNA sequencing for the discovery, mutations characterization, and expression profiling of small RNAs by leveraging of advanced NGS technologies.

Small RNA Sequencing

Small RNA species include microRNA (miRNA), small interfering RNA (siRNA), and piwi-interacting RNA (piRNA), which are a type of lowly abundant, short in length (15–200 nt), non-protein-coding RNAs that lack polyadenylation. Small RNA populations can vary significantly between different tissue types and species.

miRNAs in particular are well-studied modulators of protein expression, they are highly conserved and short pieces of RNA (typically 18-25 nt) which regulate genes at the transcriptional levels and direct post-transcriptional gene silencing of their targets by imperfect hybridization to the 3’ UTRs of mRNAs. Small RNAs have been demonstrated to be involved in a number of biological processes including development, cell proliferation and differentiation, and apoptosis.

By taking advantage of tremendous output with unprecedented sensitivity and dynamic range, NGS can identify weakly expressed small RNAs as well as quantitatively reveal heterogeneity in length and sequence, which is a powerful tool for investigating the function of small RNAs and prediction of potential mRNA target molecules. Obtaining a good small RNA-seq library begins with isolation of small RNAs by size fractionation using gel electrophoresis selection or use of silica spin columns from total RNA. Following RNA adapter ligation using a 5’ adenylated DNA adapter with a blocked 3’end, small RNAs are reverse transcribed, amplified by PCR and sequenced. To identify and annotate known miRNAs, the sequencing reads can be mapped to a species-specific database, such as miRBase. The experimental pipeline of small RNA sequencing is demonstrated in Figure 1.

Schematic  workflow of small RNA sequencing process Figure 1. Schematic workflow of small RNA sequencing process.

Sequencing Strategy and Recommended Depth

  • Illumina HiSeq SE50
  • ≥ 10 M reads

Data Analysis

We have a team with the expert knowledge and computational resources to help you achieve your data analysis objectives. Our comprehensive small RNA sequencing data analysis pipeline includes bases quality assessment, sequence length filter, small RNA distribution pattern, reference-based mapping, small RNA quantification and classification (rRNA, tRNA, miRNA, snRNA, snoRNA, tRNA, et al.), novel miRNA prediction, differential expression analysis, target gene prediction and annotation, miRNA editing analysis, miRNA family analysis, and correlation analysis for small RNA + mRNA.

Sample Requirements

  1. Sample type: Total RNA without degradation or DNA contamination.
  2. Starting amount of total RNA: ≥ 5 µg
  3. Sample conc.: ≥ 200 ng/µl
  4. Sample purity: OD260/280 = 1.8~2.2

Key Features and Advantages

  • Profile all small RNAs in any organism, of any size and quantity, whether known and unknown
  • Characterization of small RNA isoforms and their effects on gene expression
  • High resolution and high accuracy
  • Stringent quality controls
  • Dedicated support from experienced RNA and NGS specialists
  • Flexible and customizable bioinformatics analysis and data interpretation
  • Cost-effective services

By advantage of the cutting-edge Illumina platform, with the SE50 sequencing strategy and widely accepted analysis software, CD Genomics provides comprehensive solution for your needs to investigate miRNA and other types of small RNAs, including siRNA snoRNA, and piRNA. We can also help in the experimental design at the very beginning of your project to maximize the value of your data.

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