CRISPR/Cas9 Gene Editing in Bacteria: Leading to Hematopoietic Stem Cell Editing in Pakistan

Abstract

Background: Deletion of LacZ gene in E. coli HB101-pBRKan using Clustered Regularly Inter-Spaced Short Palindromic Repeats-Cas9 (CRISPR/Cas9) gene-editing tool.

Methodology: Using BioRadTM USA “Out of the blue” gene editing kit, we demonstrated in vitro knock-out model for deletion of LacZ gene in E. coli HB101-pBRKan by employing CRISPR/Cas9 gene-editing tool. We followed an already designed modular plasmid assembly strategy and tested a single nuclease. Editing genomic locus was achieved within three days of minimal lab work.

Results: We were successful in gene editing of the E. coli LacZ gene using CRISPR/Cas9.

Conclusion:  With the practical experience gained, we are confident enough to experiment in animal models to tackle the hematological disease.