Questions about the detailed experimental design for perturb-seq?

11 April 2025 1 206 Report

Hi there,

I am reading all the previous and current papers about perturb-seq and planning to carry out some sort of similar experiments very soon.

I am wondering, in the 2016 Cell paper, the lentivirus backbone encoding the single guide RNA, why the author emphasized the whole region of U6-sgRNA-EF1alpha-puro-T2A-BFP-GBC-polyA tail is anti-parallel cassette? What are the specific purposes of doing so?

Anyone can help to explain or share their experience on the Perturb-seq construction details would be of great help!

Thanks all in advance.

Lu Wei

Reza Aghanouri

Perturb-seq is a powerful technique that combines CRISPR-based genetic perturbations with single-cell RNA sequencing (scRNA-seq) to profile transcriptomic changes in individual cells following targeted gene manipulations. Any things about this are interested.

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