Multiplexed single-cell RNA-seq via transient barcoding for simultaneous expression profiling of various drug...

Area of Science:

• Genomics
• Molecular Biology
• Bioinformatics

Background:

• High-throughput single-cell RNA sequencing (scRNA-seq) offers valuable insights into cellular heterogeneity and gene expression.
• Current scRNA-seq methods face challenges in simultaneously analyzing multiple samples, limiting large-scale studies.
• The demand for scalable and efficient scRNA-seq protocols is increasing across various biological research fields.

Purpose of the Study:

• To develop and validate a universal sample barcoding method for simultaneous scRNA-seq analysis of multiple samples.
• To overcome the limitations of existing methods for multiplexing samples in scRNA-seq experiments.
• To enable cost-effective and widespread application of scRNA-seq for diverse experimental designs.

Main Methods:

• Developed a universal sample barcoding strategy using transient transfection with short barcode oligonucleotides.
• Validated the method's accuracy through a species-mixing experiment to identify multiplets and negatives.
• Applied the barcoding method to a 48-plex drug treatment experiment, followed by Drop-Seq analysis.

Main Results:

• The barcoding method accurately distinguished between different samples in a multiplexed scRNA-seq run.
• Successfully identified multiplets and negative cells, ensuring data quality.
• Revealed unique transcriptome responses and target-specific gene expression signatures for each drug in a 48-plex experiment at the single-cell level.

Conclusions:

• The developed universal sample barcoding method is accurate, cost-effective, and widely applicable for multiplexed scRNA-seq.
• This method facilitates the simultaneous analysis of multiple experimental conditions, enhancing the scalability of single-cell studies.
• Enables broader adoption of scRNA-seq for comprehensive biological investigations and drug discovery.