CRISPR-Cas Single-Guide RNA Construction for Tail Fiber Gene Gp0047 of Salmonella Phage SSE-121

Year 2025, Volume: 9 Issue: 3, 25 - 37

Erlia Narulita , Annisyah Nurmitha Oktarina , Rina B. Opulencia , Agung Haris Widianto , Riska Ayu Febrianti , Hardian Susilo Addy

Abstract

In this study, we conducted an in-silico study of single guide RNA (sgRNA) construction as the initial stage of CRISPR-Cas9-based genome editing on Salmonella bacteriophage SSE-121 to broaden its host range. The results of the study discovered 188 sgRNA candidates using CHOPCHOP prediction. All selected candidates were docked using free website-based docking tools HNADOCK, therefore the top seven candidates of sgRNAs were docked using HDOCK with the Cas9 protein. The molecular dynamics simulation of the most optimum sgRNA-Cas9 protein (docking score -387.43) and sgRNA-Cas9-DNA (docking score -431.58) were calculated using AMBER14 forcefield in YASARA Dynamics. Based on the in-silico evaluation, the gRNA6 was obtained as the optimum single guide RNA. The docking score with DNA targets was -592.23 and Cas9 protein was -387.43. The comparison between the Cas9 and Cas9-sgRNA complex showed that the binding effect of sgRNA could maintain better conformational stability than in the single form.

Keywords

CRISPR, genome editing, molecular docking, molecular dynamics

Supporting Institution

University of Jember

Project Number

3791/UN25.3.1/LT/2023

References

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