Abstract
Bacterial DNA methylation is involved in diverse cellular functions, including modulation of gene expression, DNA repair, and restriction-modification systems for defense against viruses and other foreign DNA. Restriction systems hinder efforts to engineer organisms to produce fuels and chemicals from waste and renewable feedstocks by degrading DNA during transformation. Methylome analysis allows identification of motifs within a bacterial chromosome that may be targeted by native restriction enzymes. Further expression of the corresponding methyltransferases in Escherichia coli allows plasmid DNA to be protected from restriction in the target organism, thereby drastically enhancing transformation efficiency. Nanopore sequencing can detect methylated bases, but software is needed to transform modified base coordinates into methylated motifs. Here, we develop MIJAMP (MIJAMP Is Just A MethylBED Parser), a software package that was developed to discover methylated motifs from the output of ONT's Modkit or other data in the methylBED format. MIJAMP employs a human-driven refinement strategy that empirically validates all motifs against genome-wide methylation data, thus eliminating incorrect motifs. MIJAMP also reports methylation data on specific, user-defined motifs. Using MIJAMP, we determined the methylated motifs both in a control strain (wild-type E. coli) and in Synecococcus sp. strain PCC7002, laying the foundation for improved transformation in this organism. MIJAMP is available at https://code.ornl.gov/alexander-public/mijamp/. One Sentence Summary: Here we describe software written to discover DNA methylation motifs from nanopore sequencing data.
| Original language | English |
|---|---|
| Article number | kuaf022 |
| Journal | Journal of Industrial Microbiology and Biotechnology |
| Volume | 52 |
| DOIs | |
| State | Published - 2025 |
Funding
We thank Andrew Hren at CU—Boulder for the sample of gDNA from Synecococcus sp. PCC7002. We also thank Dr. Andrea Garza Elizondo for her work on the graphical abstract and Fig. . This manuscript has been authored by Oak Ridge National Laboratory, which is managed by UT-Battelle, LLC, under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy. The United States Government retains and the publisher, by accepting the article for publication, acknowledges that the United States Government retains a non-exclusive, paid-up, irrevocable, world-wide license to publish or reproduce the published form of this manuscript, or allow others to do so, for United States Government purposes. The Department of Energy will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan ( http://energy.gov/downloads/doe-public-access-plan ). Funding was provided in part by the U.S. DOE Office of Energy Efficiency and Renewable Energy Bioenergy Technologies Office (BETO) to the Agile BioFoundry. Funding was also provided in part by the DOE Office of Science Office of Biological and Environmental Research award DE-SC0023085.
Keywords
- ONT sequencing
- methylome
- restriction systems