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However, the impact of strain selection on the outcome of pan-genome analysis is poorly understood. Furthermore, a standard protocol to ensure high-quality pan-genome results is lacking. In this study, we carried out a series of pan-genome analyses of different strain sets of Bacillus subtilis to understand the impact of various strains on the performance and output quality of pan-genome analyses. Consequently, we found that the results obtained by pan-genome analyses of B. subtilis can be influenced by the inclusion of incorrectly classified Bacillus subspecies strains, phylogenetically distinct strains, engineered genome-reduced strains, chimeric strains, strains with a large number of unique genes or a large proportion of pseudogenes, and multiple clonal strains. Since the presence of these confounding strains can seriously affect the quality and true landscape of the pan-genome, we should remove these deviations in the process of pan-genome analyses. Our study provides new insights into the removal of biases from confounding strains in pan-genome analyses at the beginning of data processing, which enables the achievement of a closer representation of a high-quality pan-genome landscape of B. subtilis that better reflects the performance and credibility of the B. subtilis pan-genome. This procedure could be added as an important quality control step in pan-genome analyses for improving the efficiency of analyses, and ultimately contributing to a better understanding of genome function, evolution and genome-reduction strategies for B. subtilis in the future.<\/jats:p>","DOI":"10.1093\/bib\/bbaa013","type":"journal-article","created":{"date-parts":[[2020,1,22]],"date-time":"2020-01-22T12:16:27Z","timestamp":1579695387000},"page":"1951-1971","source":"Crossref","is-referenced-by-count":49,"title":["Toward a high-quality pan-genome landscape of<i>Bacillus subtilis<\/i>by removal of confounding strains"],"prefix":"10.1093","volume":"22","author":[{"given":"Hao","family":"Wu","sequence":"first","affiliation":[{"name":"Department of Physics, School of Science, Tianjin University"}]},{"given":"Dan","family":"Wang","sequence":"additional","affiliation":[{"name":"Department of Physics, School of Science, Tianjin University"}]},{"given":"Feng","family":"Gao","sequence":"additional","affiliation":[{"name":"Department of Physics, School of Science, and the Frontier Science Center of Synthetic Biology (MOE), Key Laboratory of Systems Bioengineering (MOE), Tianjin University"}]}],"member":"286","published-online":{"date-parts":[[2020,2,17]]},"reference":[{"key":"2021032314275272300_ref1","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1186\/1745-6150-6-1","article-title":"The advantages and disadvantages of horizontal gene transfer and the emergence of the first species","volume":"6","author":"Vogan","year":"2011","journal-title":"Biol Direct"},{"key":"2021032314275272300_ref2","doi-asserted-by":"crossref","first-page":"6688","DOI":"10.1093\/nar\/gkn668","article-title":"Genomics of bacteria and archaea: the emerging dynamic view of the prokaryotic world","volume":"36","author":"Koonin","year":"2008","journal-title":"Nucleic Acids Res"},{"key":"2021032314275272300_ref3","doi-asserted-by":"crossref","first-page":"376","DOI":"10.1093\/embo-reports\/kve097","article-title":"Ecological fitness, genomic islands and bacterial pathogenicity. 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