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RNA-seq

RNA-seq

RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases.
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while microarray-based...

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Updated: Jun 10, 2026

Preparation of Small RNA Libraries for Sequencing from Early Mouse Embryos

Preparation of Small RNA Libraries for Sequencing from Early Mouse Embryos

Published on: October 9, 2020

Single Cell Micro RNA Sequencing Library Preparation.

Sarah M Hücker¹, Stefan Kirsch²

¹Fraunhofer Institut für Toxikologie und Experimentelle Medizin, Abteilung Personalisierte Tumortherapie, Regensburg, Germany.

Methods in Molecular Biology (Clifton, N.J.)

|January 9, 2024

Summary

This study presents a new protocol for micro RNA (miRNA) sequencing library preparation from very low input samples, including single cells. This method enables accurate miRNA quantification for biomarker discovery and disease research.

Keywords:

CTC NGS Single-cell sequencing miRNA ncRNA small RNA

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Retrospective MicroRNA Sequencing: Complementary DNA Library Preparation Protocol Using Formalin-fixed Paraffin-embedded RNA Specimens

Retrospective MicroRNA Sequencing: Complementary DNA Library Preparation Protocol Using Formalin-fixed Paraffin-embedded RNA Specimens

Published on: May 5, 2018

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Last Updated: Jun 10, 2026

Preparation of Small RNA Libraries for Sequencing from Early Mouse Embryos

Preparation of Small RNA Libraries for Sequencing from Early Mouse Embryos

Published on: October 9, 2020

Retrospective MicroRNA Sequencing: Complementary DNA Library Preparation Protocol Using Formalin-fixed Paraffin-embedded RNA Specimens

Retrospective MicroRNA Sequencing: Complementary DNA Library Preparation Protocol Using Formalin-fixed Paraffin-embedded RNA Specimens

Published on: May 5, 2018

Area of Science:

Molecular Biology
Genetics
Biochemistry

Background:

Micro RNAs (miRNAs) are key post-transcriptional regulators involved in cellular functions and disease pathogenesis.
Accurate quantification of miRNA expression is crucial for understanding their roles and for developing diagnostic and prognostic biomarkers.
Current methods face challenges in quantifying miRNAs from limited biological material, such as single cells.

Purpose of the Study:

To develop and describe a robust protocol for micro RNA sequencing library preparation from low-input samples.
To enable accurate miRNA expression profiling at the single-cell level.
To facilitate the use of miRNAs as biomarkers in clinical and basic research.

Main Methods:

The protocol involves adapter ligation to miRNAs after cell lysis, followed by exonuclease treatment to reduce adapter dimers.
Reverse transcription, amplification, and purification steps are performed to generate the miRNA sequencing library.
The method is compatible with various single-cell isolation techniques, including micromanipulation and FACS sorting.

Main Results:

The described protocol successfully prepares miRNA sequencing libraries from very low input samples, including single cells and clinical samples like circulating tumor cells.
The protocol incorporates quality control measures to ensure the selection of high-quality samples for sequencing.
This method allows for precise miRNA quantification essential for biomarker discovery.

Conclusions:

The developed protocol significantly advances the ability to profile miRNA expression from challenging low-input samples.
This technique holds promise for identifying novel miRNA biomarkers for disease diagnosis and prognosis.
It provides a valuable tool for basic research into the physiological functions of miRNAs.