Anti-RNA virus crRNA targets efficient screening platform based on bioinformatics and CRISPR detection.

Area of Science:

• Molecular Biology
• Virology
• Bioinformatics

Background:

• RNA virus mutation and evolution present significant therapeutic challenges.
• The CRISPR-Cas system offers programmable antiviral capabilities.
• Traditional cell-based screening for CRISPR targets is time-consuming and inefficient.

Purpose of the Study:

• To develop a rapid and efficient screening platform for CRISPR RNA (crRNA) targets.
• To accelerate the identification of effective antiviral crRNAs.
• To validate the platform's efficacy against SARS-CoV-2 and Dengue virus (DENV).

Main Methods:

• Combined CaSilico-based bioinformatics with CRISPR in vitro detection technology.
• Bioinformatic screening to design and characterize crRNAs and target sequences.
• In vitro CRISPR assays for secondary screening and identification of optimal cleavage efficiency.

Main Results:

• Successfully designed and screened effective crRNAs targeting conserved regions of the SARS-CoV-2 N gene.
• Demonstrated SARS-CoV-2 inhibition in HEK 293T cells.
• Developed and validated crRNAs targeting DENV, reducing viral RNA load by over 90% and inhibiting all four serotypes in mammalian cell lines.

Conclusions:

• The developed platform significantly reduces screening time for antiviral targets compared to traditional methods.
• This approach provides a novel and efficient strategy for screening antiviral crRNAs.
• The platform shows promise for developing new CRISPR-based antiviral therapies.