Endogenous CRISPR/Cas Systems Prediction: A Glimpse towards Harnessing CRISPR/ Cas Machineries for Genetic Engineering

Genetic engineering field has become an imperative approach for enhancement of various bioproducts yield and productivity; and found extended applications in gene therapy, nanotechnology, as well as industrial microbiology. Modern genetic engineering tool CRISPR/Cas system, specifically the Type II system from Streptococcus pyogenes, is gaining traction in recent years and being utilized to engineer novel strains to overproduce primary fermentation product of interest. Employing this technology for non-model microorganism such as Clostridium spp is still restricted due to several limitations such as inadequate genome information, resistance against transformation, low plasmid replication, and the ability for gene expression. The prediction of CRISPR/Cas systems in microbial genomes is fundamentally the initial step towards exploitation of this technology to engineer Clostridium spp. In this study, we demonstrate a simple yet effective method to predict component of endogenous CRISPR/Cas systems, using Clostridium spp genomes as a proof-of- concept. We identified the “real” CRISPR array together with the cas gene operon consist of Type I‑B signature proteins in Clostridium pasteurianum which is in agreement with the previous report, implying that this strategy generates reliable CRISPR/Cas systems prediction. Thus, this provides a glimpse on how bioinformatics and biocomputational tools can be utilized to overcome barriers in genetic engineering.