small RNA-seq

small RNA-seq (full name: small RNA sequencing) is a technique used to identify and analyse small RNA molecules (typically 18-30 nucleotides long), such as microRNA (miRNA), small interfering RNA (siRNA) and piRNA (piwi-interacting RNA).

Small RNA-seq workflow

Applications of small RNA-seq

This technique provides a better understanding of the role of small RNAs in cell biology and their potential therapeutic applications.

Technical specifications of small RNA-seq

• Library preparation: ribosomal RNA is removed from the sample, followed by selection of the short-RNA fraction.
• Sequencing mode: 1 x 50 bp (SE50)
• Sequencers: Illumina platforms
• Recommended amount of data per sample:
  • 10M reads per sample
  • 20M reads per sample
  • Other configurations are also available depending on the client's needs.
• Data quality: Q30 ≥ 85%
• Recommended Sample Requirements - DNA/RNA:
  • Sample type: total RNA containing a small-RNA fraction
  • Amount: > 2000 ng
  • Concentration (NanoDrop): >100 ng/μL
  • Volume: >10 μL
  • Purity (OD260/280): 1.8 - 2.2
  • RIN: >7
• MINIMUM Sample Requirements - DNA/RNA:
  • Sample type: total RNA
  • Amount: > 500 ng
  • Concentration (Qubit): >80 ng/μL
  • Volume: >10 μL
  • Purity (OD260/280): 1.7 - 2.5
  • RIN: >6.5
  • 5.0 ≥ 28S/18S ≥ 1.0

RNA isolation from the supplied samples is also available. Details here.

Small RNA-seq bioinformatics analysis

The advanced bioinformatics analysis included in the small RNA-seq obejmuje m.in.:

• Quality control of sequencing data
• Annotation of ncRNA and repetitive sequences, followed by mapping to the reference genome
• Identification of known and novel miRNAs
• Prediction of novel miRNA precursors using miRDeep2
• miRNA expression profiling
• Identification and annotation of miRNA target genes
• KEGG annotation of target genes of differentially expressed miRNAs