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Related Concept Videos

RNA-seq03:21

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...
Next-generation Sequencing03:00

Next-generation Sequencing

The first human genome sequencing project cost $2.7 billion and was declared complete in 2003, after 15 years of international cooperation and collaboration between several research teams and funding agencies. Today, with the advent of next-generation sequencing technologies, the cost and time of sequencing a human genome have dropped over 100 fold.
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features.

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Related Experiment Video

Updated: May 12, 2026

Multiplexed Single Cell mRNA Sequencing Analysis of Mouse Embryonic Cells
08:30

Multiplexed Single Cell mRNA Sequencing Analysis of Mouse Embryonic Cells

Published on: January 7, 2020

An Advanced Single-Cell RNA Sequencing (scRNA-seq) Protocol Utilizing Custom-Designed Multiplexing.

Feng Gao1, Xujie Liu1,2, Fan Sun2

  • 1Department of Immunology and Immune Therapeutics, Hastings Center for Pulmonary Research, Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, Los Angeles, CA, USA.

Bio-Protocol
|May 11, 2026
PubMed
Summary
This summary is machine-generated.

A new biotin-streptavidin method enables reliable cell hashing for single-cell RNA sequencing in mouse strains where commercial kits fail. This approach improves multiplexing efficiency and data quality for diverse research applications.

Keywords:
5′ gene expressionBiotin-streptavidin conjugationCell hashingFVB/N miceMHC-I haplotype mismatchSample multiplexingscRNA-seq

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Last Updated: May 12, 2026

Multiplexed Single Cell mRNA Sequencing Analysis of Mouse Embryonic Cells
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AQRNA-seq for Quantifying Small RNAs
05:12

AQRNA-seq for Quantifying Small RNAs

Published on: February 2, 2024

Area of Science:

  • Single-cell genomics
  • Immunology
  • Molecular biology

Background:

  • Cell hashing using single-cell RNA sequencing (scRNA-seq) improves experimental efficiency by minimizing batch effects.
  • Commercial mouse hashtags frequently fail in specific strains like FVB/N due to antibody-epitope incompatibilities, particularly with MHC-I.
  • This limitation hinders strain-specific or custom multiplexing strategies in single-cell transcriptomics.

Purpose of the Study:

  • To develop and validate a robust, strain-compatible cell hashing method for scRNA-seq.
  • To overcome the limitations of commercial hashtags in FVB/N and other mouse strains.
  • To provide a customizable multiplexing solution for single-cell transcriptomics.

Main Methods:

  • Utilized biotinylated antibody cocktails and streptavidin-conjugated oligos for sample multiplexing.
  • Integrated the biotin-streptavidin approach with the 10× Genomics 5' single-cell gene expression and V(D)J profiling workflow.
  • Incorporated anti-CD326 (Ep-CAM) for enhanced labeling of non-immune cells like epithelial and tumor cells.

Main Results:

  • Successfully validated the biotin-streptavidin cell hashing protocol in FVB/N mouse lung tissues.
  • Achieved high-quality single-cell libraries with reliable sample multiplexing.
  • Demonstrated compatibility with multi-omic 5' workflows, enabling joint gene expression and V(D)J repertoire profiling.

Conclusions:

  • The biotin-streptavidin method offers a practical and effective alternative for cell hashing in mouse strains incompatible with commercial kits.
  • This protocol supports strain-specific and custom multiplexing strategies, enhancing the utility of scRNA-seq.
  • The customizable framework allows adaptation for various mouse strains and tissue-specific cell markers, broadening its applicability.