PRODUCT

CRISPR Guide Enrichment Kit

Amplify a CRISPR guide sequence derived from a CROP-style vector alongside your standard RNA Sequencing Library.

Description

The ScaleBio CRISPR Guide Enrichment Kit is modified from our standard ScaleBio Single Cell RNA Sequencing Kit, with the added benefit of capturing a CRISPR guide sequence derived from a CROP-style vector.

To maximize guide detection, we have included target-specific priming during reverse transcription, as well as PCR enrichment in the final step of library preparation.

This kit enables the processing of up to 500,000 cells or nuclei per run with extended throughput, as well as the multiplexing of up to 96 samples.

Specifications

Cell Throughput

125,000 cells or nuclei per run, or up to 500,000 cells or nuclei per run with extended throughput

Recommended input

10,000 cells per well (Initial Distribution)

Sample Throughput

Up to 96 samples per run

CRISPR Guide Enrichment Workflow

CRISPR libraries are designed, assembled into lentiviral vectors, which are used to transfect cells with individual guides.

Cells are then fixed according to the ScaleBio Sample Fixation Protocol. A guide-specific RT primer is included during Reverse Transcription indexing, improving guide detection. From the final pooled library, a guide enrichment PCR is performed prior to sequencing.

30% increase in guide detection with ScaleBio's custom RT Primer

Human cells were used to test guide detection using only PolyA priming during reverse transcription vs. using the custom RT primer in the ScaleBio CRISPR Guide Enrichment Kit. The use of a custom RT primer significantly improves guide detection and thereby enables lower overall sequencing costs.

Custom Priming substantially increases guide detection rate across cell types

When comparing guide detection rates (≥3 UMIs per guide) using a traditional CROP-seq primer, a higher percentage of guides are detected both at the population level as well as in the percentage of cells with an identified guide with the ScaleBio CRISPR Guide Enrichment Kit.