Biology subjects

Petrov, V. A.

Publications and source records attributed to Petrov, V. A..

3 recordsLinked to original sources

bioRxiv2023

CRISPR-COPIES: An in silico platform for discovery of neutral integration sites for CRISPR/Cas-facilitated gene integration

The CRISPR/Cas system has emerged as a powerful tool for genome editing in metabolic engineering and human gene therapy. However, locating the optimal site on the chromosome to integrate heterologous genes using the CRISPR/Cas system remains an open question. Selecting a suitable site for gene integration involves considering multiple complex criteria, including factors related to CRISPR/Cas-mediated integration, genetic stability, and gene expression. Consequently, identifying such sites on specific or different chromosomal locations typically requires extensive characterization efforts. To address these challenges, we have developed CRISPR-COPIES, a COmputational Pipeline for the Identification of CRISPR/Cas-facilitated intEgration Sites. This tool leverages ScaNN, a state-of-the-art model on the embedding-based nearest neighbor search for fast and accurate off-target search and can identify genome-wide intergenic sites for most bacterial and fungal genomes within minutes. As a proof of concept, we utilized CRISPR-COPIES to characterize neutral integration sites in three diverse species: Saccharomyces cerevisiae, Cupriavidus necator, and a human cell line. In addition, we developed a user-friendly web interface for CRISPR-COPIES (https://biofoundry.web.illinois.edu/copies/). We anticipate that CRISPR-COPIES will serve as a valuable tool for targeted DNA integration and aid in the characterization of synthetic biology toolkits, enable rapid strain construction to produce valuable biochemicals and support human gene and cell therapy applications. Graphical abstractOverview and application of CRISPR-COPIES in the field of biotechnology. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=93 SRC="FIGDIR/small/556564v1_ufig1.gif" ALT="Figure 1"> View larger version (20K): org.highwire.dtl.DTLVardef@c723a1org.highwire.dtl.DTLVardef@1f42594org.highwire.dtl.DTLVardef@1cf4bd6org.highwire.dtl.DTLVardef@7c1028_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2022

PlasmidMaker: a Versatile, Automated, and High Throughput End-to-End Platform for Plasmid Construction

Plasmids are used extensively in basic and applied biology. However, design and construction of plasmids, specifically the ones carrying complex genetic information, remains one of the most time-consuming, labor-intensive, and rate-limiting steps in performing sophisticated biological experiments. Here, we report the development of a versatile, robust, automated end-to-end platform named PlasmidMaker that allows error-free construction of plasmids with virtually any sequences in a high-throughput manner. This platform consists of a most versatile DNA assembly method using Pyrococcus furiosus Argonaute (PfAgo)-based artificial restriction enzymes, a user-friendly frontend for plasmid design, and a backend that streamlines the workflow and integration with a robotic system. As a proof of concept, we used this platform to generate 101 plasmids from six different species ranging from 5 to 18 kb in size from up to 11 DNA fragments within 3 days. PlasmidMaker should greatly expand the potential of synthetic biology.

synthetic biology↗

bioRxiv2018

Biliary microbiota and bile acids composition in cholelithiasis

BackgroundA functional interplay between BAs and microbial composition in gut is a well-documented phenomenon. In bile this phenomenon is far less studied and with this report we describe the interactions between the BAs and microbiota in this complex biological matrix.\n\nMethodologyThirty-seven gallstone disease patients of which twenty-one with Opisthorchis felineus infection were enrolled in the study. The bile samples were obtained during laparoscopic cholecystectomy for gallstone disease operative treatment. Common bile acids composition were measured by LC-MS/MS using a column in reverse phase. For all patients gallbladder microbiota was previously analyzed with 16S rRNA gene sequencing on Illumina MiSeq platform. The associations between bile acids composition and microbiota were analysed.\n\nPrincipal findingsBile acids signature and O. felineus infection status exerts influence on beta-diversity of bile microbial community. Direct correlations were found between taurocholic acid, taurochenodeoxycholic acid concentrations and alpha-diversity of bile microbiota. Taurocholic acid and taurochenodeoxycholic acid both shows positive associations with the presence of Chitinophagaceae family, Microbacterium and Lutibacterium genera and Prevotella intermedia. Also direct associations were identified for taurocholic acid concentration and the presence of Actinomycetales and Bacteroidales orders, Lautropia genus, Jeotgalicoccus psychrophilus and Haemophilus parainfluenzae as well as for taurochenodeoxycholic acid and Acetobacteraceae family and Sphingomonas genus. There were no differences in bile acids concentrations between O. felineus infected and non-infected patients.\n\nConclusions/SignificanceAssociations between diversity, taxonomic profile of bile microbiota and bile acids levels were evidenced in patients with cholelithiasis. Increase of taurochenodeoxycholic acid and taurocholic acid concentration correlates with bile microbiota alpha-diversity and appearance of opportunistic pathogens. Alteration of bile acids signature could cause shifts in bile microbial community structure.

microbiology↗