LOCUS       sequence-207569-        4559 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transposon mutagenesis and Tn-seq.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4559)
  AUTHORS   Wiles TJ, Norton JP, Russell CW, Dalley BK, Fischer KF, Mulvey MA
  TITLE     Combining quantitative genetic footprinting and trait enrichment 
            analysis to identify fitness determinants of a bacterial pathogen.
  JOURNAL   PLoS Genet. 2013;9(8):e1003716. doi: 10.1371/journal.pgen.1003716. 
            Epub 2013 Aug 22.
  PUBMED    23990803
REFERENCE   2  (bases 1 to 4559)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS 
            Genet."; date: "2013"; volume: "9(8):e1003716. doi"; pages: " 
            10.1371/journal.pgen.1003716. Epub 2013 Aug 22"
COMMENT     Sequence Label: pSAM_Ec
FEATURES             Location/Qualifiers
     source          1..4559
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..22
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(37..42,43..60,61..67)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     rep_origin      1170..1558
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     oriT            1673..1782
                     /note="incP origin of transfer"
     CDS             complement(join(1946..2737,2738..2806))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2569..2588
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2807..2911)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2979..2997
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             3404..4198
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3458..3477)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4068..4087
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     terminator      4257..4343
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4435..4462
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
       61 tatgttgtgt ggaattgtga gcggatacat atggaaaaaa aggaatttcg tgttttgata
      121 aaatactgtt ttctgaaggg aaaaaataca gtggaagcaa aaacttggct tgataatgag
      181 tttccggact ctgccccagg gaaatcaaca ataattgatt ggtatgcaaa attcaagcgt
      241 ggtgaaatga gcacggagga cggtgaacgc agtggacgcc cgaaagaggt ggttaccgac
      301 gaaaacatca aaaaaatcca caaaatgatt ttgaatgacc gtaaaatgaa gttgatcgag
      361 atagcagagg ccttaaagat atcaaaggaa cgtgttggtc atatcattca tcaatatttg
      421 gatatgcgga agctctgtgc gaaatgggtg ccgcgcgagc tcacatttga ccaaaaacaa
      481 cgacgtgttg atgattctaa gcggtgtttg cagctgttaa ctcgtaatac acccgagttt
      541 ttccgtcgat atgtgacaat ggatgaaaca tggctccatc actacactcc tgagtccaat
      601 cgacagtcgg ctgagtggac agcgaccggt gaaccgtctc cgaagcgtgg aaagactcaa
      661 aagtccgctg gcaaagtaat ggcctctgtt ttttgggatg cgcatggaat aatttttatc
      721 gattatcttg agaagggaaa aaccatcaac agtgactatt atatggcgtt attggagcgt
      781 ttgaaggtcg aaatcgcggc aaaacggccc cacatgaaga agaaaaaagt gttgttccac
      841 caagacaacg caccgtgcca caagtcattg agaacgatgg caaaaattca tgaattgggc
      901 ttcgaattgc ttccccaccc gccgtattct ccagatctgg cccccagcga ctttttcttg
      961 ttctcagacc tcaaaaggat gctcgcaggg aaaaaatttg gctgcaatga agaggtgatc
     1021 gccgaaactg aggcctattt tgaggcaaaa ccgaaggagt actaccaaaa tggtatcaaa
     1081 aaattggaag gtcgttataa tcgttgtatc gctcttgaag ggaactatgt tgaataagcg
     1141 gccgccaccg cggtggaggg gaattcccat gtcagccgtt aagtgttcct gtgtcactca
     1201 aaattgcttt gagaggctct aagggcttct cagtgcgtta catccctggc ttgttgtcca
     1261 caaccgttaa accttaaaag ctttaaaagc cttatatatt cttttttttc ttataaaact
     1321 taaaacctta gaggctattt aagttgctga tttatattaa ttttattgtt caaacatgag
     1381 agcttagtac gtgaaacatg agagcttagt acgttagcca tgagagctta gtacgttagc
     1441 catgagggtt tagttcgtta aacatgagag cttagtacgt taaacatgag agcttagtac
     1501 gtgaaacatg agagcttagt acgtactatc aacaggttga actgctgatc ttcagatcct
     1561 ctacgccgga cgcatcgtgg ccggatcaat tccgtgatag gtgggctgcc cttcctggtt
     1621 ggcttggttt catcagccat ccgcttgccc tcatctgtta cgccggcggt agccggccag
     1681 cctcgcagag caggattccc gttgagcacc gccaggtgcg aataagggac agtgaagaag
     1741 gaacacccgc tcgcgggtgg gcctacttca cctatcctgc ccggctgacg ccgttggata
     1801 caccaaggaa agtctacacg aaccctttgg caaaatcctg tatatcgtgc gaaaaaggat
     1861 ggatataccg aaaaaatcgc tataatgacc ccgaagcagg gttatgcagc ggaaaacgga
     1921 attgatcccc ctcgaattga cgcgtttacc aatgcttaat cagtgaggca cctatctcag
     1981 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     2041 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     2101 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     2161 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     2221 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     2281 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     2341 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     2401 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     2461 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     2521 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     2581 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     2641 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     2701 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     2761 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     2821 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     2881 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     2941 tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct
     3001 ttcgtcacgc gtcctcggta cctaacaggt tggatgataa gtccccggtc ttcgtatgcc
     3061 gtcttctgct tggcgcgccc tcgagcaatt gaggaagcgg aacacgtaga aagccagtcc
     3121 gcagaaacgg tgctgacccc ggatgaatgt cagctactgg gctatctgga caagggaaaa
     3181 cgcaagcgca aagagaaagc aggtagcttg cagtgggctt acatggcgat agctagactg
     3241 ggcggtttta tggacagcaa gcgaaccgga attgccagct ggggcgccct ctggtaaggt
     3301 tgggaagccc tgcaaagtaa actggatggc tttcttgccg ccaaggatct gatggcgcag
     3361 gggatcaaga tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg
     3421 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca
     3481 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt
     3541 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg
     3601 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga
     3661 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca
     3721 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct
     3781 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac
     3841 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc
     3901 gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt
     3961 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt
     4021 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg
     4081 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat
     4141 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc
     4201 gggactctgg ggttcgaaat gactctagaa tgcgagagta gggaactgcc aggcatcaaa
     4261 taaaacgaaa ggctcagtcg aaagactggg cctttcgttt tatctgttgt ttgtcggtga
     4321 acgctctcct gagtaggaca aatccgccgg gagcggattt gaacgttgcg aagcaacggc
     4381 ccggagggtg gcgggcagga cgcccgccat aaactgccag gcatcaaatt aagcagaagg
     4441 ccatcctgac ggatggcctt tttgcgtttc tacctgcagg gcgcgccaag cagaagacgg
     4501 catacgaaga ccggggactt atcatccaac ctgttaggat ccagtgagcg caacgcaat
//
