CrisprBuildr: an open-source application for CRISPR-mediated genome engineering in Drosophila melanogaster

Area of Science:

• Molecular Biology
• Genomics
• Biotechnology

Background:

• CRISPR/Cas9 technology enables targeted genome engineering for gene modification and protein studies.
• Designing and cloning constructs for CRISPR/Cas9 experiments is often time-consuming and labor-intensive.
• Efficient tools are needed to streamline the process of gene deletion and protein tagging.

Purpose of the Study:

• To develop CrisprBuildr, a web-based application to simplify CRISPR/Cas9-based genome engineering.
• To facilitate the deletion of genes or insertion of fluorescent proteins (e.g., EGFP, mCherry) at specific protein termini.
• To provide a user-friendly platform for designing guide RNAs and repair template vectors.

Main Methods:

• Development of CrisprBuildr, an open-source, web-based application.
• Integration of the Drosophila melanogaster genome as a primary template, adaptable for other genomes.
• Generation of novel tagging vectors incorporating fluorescent proteins and the SspB-Q73R peptide for iLID-based optogenetics.

Main Results:

• CrisprBuildr enables users to design constructs for gene deletion or N-/C-terminal fluorescent protein insertion.
• The application guides users through the design of guide RNAs and repair template vectors.
• New tagging vectors for EGFP, mCherry, and optogenetic applications have been generated.

Conclusions:

• CrisprBuildr significantly reduces the time and effort required for CRISPR/Cas9 construct design.
• The tool supports efficient protein labeling and gene manipulation for in vivo studies.
• Future development will expand CrisprBuildr's capabilities with more vectors, genomes, and CRISPR applications.