title: bacteria
design: DenovoassemblywasperformedtogeneratesequencesofUTNGt21AsampleusingIlluminaHiSeqXTenplatform.Inbrief,thesequencinglibrarywaspreparedbyrandomfragmentationoftheDNAorcDNAsample,followedby5and3adapterligationaccordingtotheprotocolprovidedbythemanufacturer(MacrogenInc.,Seoul,Korea).Sequencingdatawereconvertedintorawdataforanalysis.TheoverallqualityofreadsgeneratedbyFastQC(v0.11.5,http://www.bioinformatics.babraham.ac.uk/projects/fastqc),totalbases,totalreads,GCcontent,andbasicstatisticswerecalculated.Toreducebiasesintheanalysis,adaptertrimmingandqualityfilteringwereperformed.Trimmomaticv0.36(http://www.usadellab.org/cms/?page=trimmomatic)wasusedtoremoveadaptersequences.Thequalityoffilteredreads,totalbases,totalreads,GCcontent,andbasicstatisticswerecalculated.DenovoassemblywasperformedbyvariouskmerusingSPAdes3.15.1(http://cab.spbu.ru/software/spades/).Kmeranalysiswasperformedtoprovideinformationaboutcoverage,heterozygosity,andestimatedgenomesize(Jellyfishv2.2.10,http://www.genome.umd.edu/jellyfish.html).Usingfilteredreads,denovoassemblywasperformedusingaDeBruijngraphassembler(http://qb.cshl.edu/genomescope/).ThereadsweresplitintomultipleKmers(OligonucleotideswithlengthK,Kequalinteger),whichwerefurtheralignedwitheachother,andthesequenceswereextendedonebaseatatimebasedontheoverlapinformationofK-mers. TheassembledgenomewasvalidatedusingmappingstrategyandBUSCOanalysis(https://busco.ezlab.org/,BUSCOversion3.0)(Seppeyetal.,2019).Thefilteredreadswerealignedtotheassembledgenome,andtheirinsertsizewasestimatedforvalidation.Toassessthecompletenessofthegenomeassembly,BUSCOanalysiswasperformedbasedonevolutionarilyinformedexpectationsofgenecontentfromnear-universalsingle copyorthologs.Bydefault,bacteriaDBwasusedforanalysis.Afterthecompletegenomeordraftgenomewasassembled,BLASTanalysiswascarriedouttoidentifytowhichspecieseachscaffoldshowedsimilarity.