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However, metagenome-assembled genomes (MAGs) are often fragmented, primarily when sequencing was performed with short reads. Recent long-read sequencing technologies promise to improve genome reconstruction. However, the integration of two different sequencing modalities makes downstream analyses complex. We, therefore, developed MUFFIN, a complete metagenomic workflow that uses short and long reads to produce high-quality bins and their annotations. The workflow is written by using Nextflow, a workflow orchestration software, to achieve high reproducibility and fast and straightforward use. This workflow also produces the taxonomic classification and KEGG pathways of the bins and can be further used for quantification and annotation by providing RNA-Seq data (optionally). We tested the workflow using twenty biogas reactor samples and assessed the capacity of MUFFIN to process and output relevant files needed to analyze the microbial community and their function. MUFFIN produces functional pathway predictions and, if provided\n                    <jats:italic>de novo<\/jats:italic>\n                    metatranscript annotations across the metagenomic sample and for each bin. MUFFIN is available on github under GNUv3 licence:\n                    <jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" ext-link-type=\"uri\" xlink:href=\"https:\/\/github.com\/RVanDamme\/MUFFIN\" xlink:type=\"simple\">https:\/\/github.com\/RVanDamme\/MUFFIN<\/jats:ext-link>\n                    .\n                  <\/jats:p>","DOI":"10.1371\/journal.pcbi.1008716","type":"journal-article","created":{"date-parts":[[2021,2,9]],"date-time":"2021-02-09T21:45:49Z","timestamp":1612907149000},"page":"e1008716","update-policy":"https:\/\/doi.org\/10.1371\/journal.pcbi.corrections_policy","source":"Crossref","is-referenced-by-count":47,"title":["Metagenomics workflow for hybrid assembly, differential coverage binning, metatranscriptomics and pathway analysis (MUFFIN)"],"prefix":"10.1371","volume":"17","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-7909-4868","authenticated-orcid":true,"given":"Renaud","family":"Van Damme","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7090-8717","authenticated-orcid":true,"given":"Martin","family":"H\u00f6lzer","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8970-5204","authenticated-orcid":true,"given":"Adrian","family":"Viehweger","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0030-7710","authenticated-orcid":true,"given":"Bettina","family":"M\u00fcller","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1947-8288","authenticated-orcid":true,"given":"Erik","family":"Bongcam-Rudloff","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7199-3957","authenticated-orcid":true,"given":"Christian","family":"Brandt","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2021,2,9]]},"reference":[{"key":"pcbi.1008716.ref001","doi-asserted-by":"crossref","first-page":"R245","DOI":"10.1016\/S1074-5521(98)90108-9","article-title":"Molecular biological access to the chemistry of unknown soil microbes: a new frontier for natural products","volume":"5","author":"J Handelsman","year":"1998","journal-title":"Chem Biol"},{"key":"pcbi.1008716.ref002","doi-asserted-by":"crossref","first-page":"47","DOI":"10.1186\/s13099-019-0331-8","article-title":"Metagenomics: aid to combat antimicrobial resistance in diarrhea","volume":"11","author":"R De","year":"2019","journal-title":"Gut Pathog"},{"key":"pcbi.1008716.ref003","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1007\/s13205-020-2057-1","article-title":"Metatranscriptomics: an approach for retrieving novel eukaryotic genes from polluted and related environments","volume":"10","author":"A Mukherjee","year":"2020","journal-title":"3 Biotech"},{"key":"pcbi.1008716.ref004","first-page":"S2","article-title":"Linking metagenomics to aquatic microbial ecology and biogeochemical cycles","volume":"65","author":"H-P Grossart","year":"2020","journal-title":"Limnol Oceanogr"},{"key":"pcbi.1008716.ref005","doi-asserted-by":"crossref","first-page":"9379","DOI":"10.1007\/s00253-019-10052-5","article-title":"Metagenomic approaches: effective tools for monitoring the structure and functionality of microbiomes in anaerobic digestion systems","volume":"103","author":"A Carabeo-P\u00e9rez","year":"2019","journal-title":"Appl Microbiol Biotechnol"},{"key":"pcbi.1008716.ref006","doi-asserted-by":"crossref","first-page":"4000","DOI":"10.1111\/1462-2920.15186","article-title":"Inclusion of Oxford Nanopore long reads improves all microbial and viral metagenome-assembled genomes from a complex aquifer system","volume":"22","author":"WA Overholt","year":"2020","journal-title":"Environ Microbiol"},{"key":"pcbi.1008716.ref007","article-title":"Assembly-free single-molecule nanopore sequencing recovers complete virus genomes from natural microbial communities","year":"2020","journal-title":"bioRxiv"},{"key":"pcbi.1008716.ref008","author":"KA Wetterstrand","year":"2020","journal-title":"DNA Sequencing Costs: Data. 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Anaconda | The World\u2019s Most Popular Data Science Platform. In: https:\/\/anaconda.com [Internet]. 5 Feb 2020 [cited 5 Feb 2020]. Available: https:\/\/www.anaconda.com\/"},{"key":"pcbi.1008716.ref023","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1145\/2723872.2723882","article-title":"An introduction to Docker for reproducible research","volume":"49","author":"C Boettiger","year":"2015","journal-title":"ACM SIGOPS Oper Syst Rev"},{"key":"pcbi.1008716.ref024","first-page":"31","article-title":"Genome sequences of rare, uncultured bacteria obtained by differential coverage binning of multiple metagenomes","author":"M Albertsen","year":"2013","journal-title":"Nat Biotechnol"},{"key":"pcbi.1008716.ref025","doi-asserted-by":"crossref","first-page":"i884","DOI":"10.1093\/bioinformatics\/bty560","article-title":"fastp: an ultra-fast all-in-one FASTQ preprocessor","volume":"34","author":"S Chen","year":"2018","journal-title":"Bioinformatics"},{"key":"pcbi.1008716.ref026","unstructured":"Wick R. rrwick\/Filtlong. 2020. 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