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This protocol identifies transposon integration events in genomes, suggesting improved annotation methods.
This protocol uses the command line tools provided by the Python package TnAtlas to identify and annotate transposon integration events in genomes. Given a set of sequencing reads, transposon sequences and genomes, the protocol looks for positions in the genomes where the transposon sequences have been integrated, annotates the sequencing reads with corresponding features of the genome, and presents a summary of the results as a spreadsheet. Integrations are identified by aligning reads to transposon sequences using NCBI's BLASTN, and annotations are made from reference genomes in GENBANK format. We have used this protocol in our group (https://biocomputationlab.com) to identify the location and gene locus of transposon inserts in the genome of Pseudomonas putida KT2440. However, this script can be used for other genomes for which the genome sequence and annotation are available. This updated version of the protocol is the first to be supported by the Python package TnAtlas, which contains library code which can be used to run the protocol programmatically (in a Python script or notebook), as well as compute and present other statistics about the results themselves. The package TnAtlas requires Python versions greater than or equal to 3.8. The protocol requires at least blastn version greater than or equal to 2.12. Optionally, the protocol also requires sickle version 1.33 or greater, and fastqc (any version), in order to perform the sequence trimming and quality control reporting. This is a description of the LAP entry LAPu-InsertsGenAnnotation-2.0.0 located in the LAP repository, specifically LAPu-InsertsGenAnnotation-3.0.0 and Github Entry TnAtlas, 2 places that you can download directly the tool used and usage examples. The major changes from previous version are: Now shipped as a Python package: The code and tools are packaged and can be installed with Python PIP. New tool for metadata annotation: The old --summary-map system has been replaced with a dedicated tool `tnmeta`, for general metadata annotation of the results. Command line options for saving intermediate files: By default, the tools create far fewer files. The options, --sam, --trim-save, --transposon-save, and --genome-save control the creation of intermediate files. Method of genome alignment: Alignment to the genome is now based on aligning the reads after the transposon sequence that has been found has been removed, giving preference to alignments earlier in the read, and also avoiding position errors of a few base pairs that can affect sequence logo results. Generates annotated genbank sequences: tnfind generates genbank files for each read that includes annotations of the transposon sequence and of the corresponding features from its position in the genome. Annotations from genbank: Annotations come from genbank reference genomes instead of separate CSV files. Multiple genomes: `tnfind` can search through multiple genome sequences at once by passing either passing multiple records in a genbank file, or multiple genbank files, to the `-genome` option.
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Grozinger et al. (2023) studied this question.
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