Genome Engineering of Primary Human B Cells Using CRISPR/Cas9

作者： Kanut Laoharawee1,2,3, Matthew J. Johnson1,2,3, Walker S. Lahr1,2,3, Joseph J. Peterson1,2,3, Beau R. Webber1,2,3, Branden S. Moriarity1,2,3 1Department of Pediatrics,University of Minnesota, 2Center for Genomic Engineering,University of Minnesota, 3Masonic Cancer Center,University of Minnesota

简介：

Overview

This study presents a detailed protocol for CRISPR/Cas9-based genome engineering in primary human B cells, enabling gene knockout and knock-in to explore gene functions and develop B-cell therapeutics.

Key Study Components

Research Area

• Genomics
• Cell Biology
• Therapeutic Development

Background

• Importance of studying gene function in B cells
• CRISPR/Cas9 technology enabling precise edits
• Potential applications in infectious disease and enzymopathies

Methods Used

• CRISPR/Cas9 transfection of primary B cells
• Primary human B cells as the biological model
• Electroporation for delivering gene editing tools

Main Results

• Successful knockout of the CD19 gene with over 95% reduction in expression
• High rates of indel formation at the CD19 locus confirmed by genomic sequencing
• Capable of introducing transgenes and editing base pairs for functional studies

Conclusions

• This protocol effectively enhances the study of B-cell biology and therapeutics.
• It provides a robust platform for gene editing, with significant implications for medical research.

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