LOCUS       V003641                 6537 bp    DNA     circular SYN 18-DEC-2018
DEFINITION  Exported.
ACCESSION   V003641
VERSION     V003641
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 6537)
  AUTHORS   Plaut RD, Stibitz S.
  TITLE     Improvements to a Markerless Allelic Exchange System for Bacillus
            anthracis
  JOURNAL   PLoS ONE 10 (12), E0142758 (2015)
   PUBMED   26624016
REFERENCE   2  (bases 1 to 6537)
  AUTHORS   Plaut RD, Stibitz S.
  TITLE     Direct Submission
  JOURNAL   Submitted (24-SEP-2015) Center for Biologics Evaluation and
            Research, OVRR, DBPAP, Food and Drug Administration, 10903 New
            Hampshire Ave, Bldg 52/72, Rm 3310, Silver Spring, MD 20993, USA
REFERENCE   3  (bases 1 to 6537)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6537)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing
            ##Assembly-Data-END##
            SGRef: number: 1; type: "Journal Article"; journalName: "PLoS ONE";
            date: "2015"; volume: "10"; issue: "12"; pages: "E0142758"
            SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
            (24-SEP-2015) Center for Biologics Evaluation and Research, OVRR,
            DBPAP, Food and Drug Administration, 10903 New Hampshire Ave, Bldg
            52/72, Rm 3310, Silver Spring, MD 20993, USA"
            SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6537
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    57..74
                     /label="I-SceI site"
                     /note="I-SceI site"
     oriT            225..333
                     /label="oriT"
                     /note="incP origin of transfer"
     promoter        complement(348..366)
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(376..392)
                     /label="M13 fwd"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     rep_origin      954..2276
                     /label="ts rep from pJRS233, pWV01"
                     /note="ts rep from pJRS233, pWV01"
     CDS             complement(954..1103)
                     /codon_start=1
                     /gene="orfD"
                     /product="OrfD"
                     /label="orfD"
                     /protein_id="ALQ43911.1"
                     /translation="MTNKEKELFAENEELKKEIKDLKERIERYREMEVELSTTIDLLRG
                     GIIE"
     gene            complement(954..1103)
                     /gene="orfD"
                     /label="orfD"
     gene            complement(1100..1798)
                     /gene="repA"
                     /label="repA"
     misc_feature    1623..1640
                     /note="mutations in RepA leading to temperature
                     sensitivity"
     CDS             complement(1865..2026)
                     /codon_start=1
                     /gene="orfC"
                     /product="orfC"
                     /label="orfC"
                     /protein_id="ALQ43915.1"
                     /translation="MVISESKKRVMISLTKEQDKKLTDMAKQKGFSKSAVAALAIEEYA
                     RKESEQKK"
     gene            complement(1865..2026)
                     /gene="orfC"
                     /label="orfC"
     CDS             complement(2067..2276)
                     /codon_start=1
                     /gene="orfB"
                     /product="orfB"
                     /label="orfB"
                     /protein_id="ALQ43916.1"
                     /translation="MGGKEANFASVLRPPIKCRVPIFVPKTLYPNWLKGLRGFSIANES
                     PTFSPTFFINLYLSSFIVVFMITK"
     gene            complement(2067..2276)
                     /gene="orfB"
                     /label="orfB"
     CDS             2410..2421
                     /label="Factor Xa site"
                     /note="Factor Xa recognition and cleavage site"
     promoter        2596..2625
                     /label="trc promoter"
                     /note="strong E. coli promoter; hybrid between the trp and
                     lac UV5 promoters"
     regulatory      2596..2601
                     /label="Ptrc"
                     /note="Ptrc"
                     /regulatory_class="minus_35_signal"
     regulatory      2619..2623
                     /label="Ptrc"
                     /note="Ptrc"
                     /regulatory_class="minus_10_signal"
     regulatory      2639..2648
                     /regulatory_class="ribosome_binding_site"
     CDS             2656..3435
                     /gene="ant1"
                     /label="Spectinomycin 9-adenylyltransferase"
                     /note="Spectinomycin 9-adenylyltransferase from
                     Staphylococcus aureus (strain N315). Accession#: P0A0D1"
     regulatory      3464..3469
                     /label="PrrnB"
                     /note="PrrnB"
                     /regulatory_class="minus_35_signal"
     regulatory      3487..3492
                     /label="PrrnB"
                     /note="PrrnB"
                     /regulatory_class="minus_10_signal"
     regulatory      3512..3517
                     /label="PFP2"
                     /note="PFP2"
                     /regulatory_class="minus_35_signal"
     regulatory      3535..3540
                     /label="PFP2"
                     /note="PFP2"
                     /regulatory_class="minus_10_signal"
     regulatory      3553..3562
                     /regulatory_class="ribosome_binding_site"
     CDS             3589..4281
                     /label="TurboRFP"
                     /note="red fluorescent protein from Entacmaea quadricolor"
     CDS             complement(4812..5759)
                     /label="Rep101"
                     /note="RepA protein needed for replication with the pSC101
                     origin"
     regulatory      complement(5790..5795)
                     /regulatory_class="minus_10_signal"
     rep_origin      complement(5807..6029)
                     /direction=LEFT
                     /label="pSC101 ori"
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     misc_feature    complement(6140..6508)
                     /label="par"
                     /note="par"
ORIGIN
        1 gcggccggag acctgtacac ccgggacgcg tggatccgtc gacatgcatg gtaccaatta
       61 ccctgttatc cctatagtac cggatcgcgc ttttccgctg cataaccctg cttcggggtc
      121 attatagcga ttttttcggt atatccatcc tttttcgcac gatatacagg attttgccaa
      181 agggttcgtg tagactttcc ttggtgtatc caacggcgtc agccgggcag gataggtgaa
      241 gtaggcccac ccgcgagcgg gtgttccttc ttcactgtcc cttattcgca cctggcggtg
      301 ctcaacggga atcctgctct gcgaggctgg ccgcgtaccc aattcgccct atagtgagtc
      361 gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt
      421 tacccaactt aatcgccttg cagcacatcc ccctttcgcc aggggataga ctgtaacatt
      481 ctcacgcata aaatcccctt tcattttcta atgtaaatct attaccttat tattaattca
      541 attcgctcat aattaatcct ttttcttatt acgcaaaatg gcccgattta agcacaccct
      601 ttattccgtt aatgcgccat gacagccatg ataattacta atactaggag aagttaataa
      661 atacgtaacc aacatgatta acaattatta gaggtcatcg ttcaaaatgg tatgcgtttt
      721 gacacatcca ctatatatcc gtgtcgttct gtccactcct gaatcccatt ccagaaattc
      781 tctagcgatt ccagaagttt ctcagagtcg gaaagttgac cagacattac gaactggcac
      841 agatggtcat aacctgaagg aagatcattt tgtttattgc aattgtattg ctattaatcg
      901 caacatcaaa ccaaaataaa aacccccttc gactttcgtc agggggcttt tatttattca
      961 ataatccctc ctctcaataa atctattgtt gtacttaatt caacttccat ttctctgtat
     1021 ctttcaatac gctcttttaa gtccttaatt tcttttttta attcctcatt ttcagcaaat
     1081 aactcttttt ctttgtttgt cattttattt cccccgtttc agcatcaaga acctttgcat
     1141 aacttgctct atatccacac tgataattgc cctcaaacca taatctaaag gcgctagagt
     1201 ttgttgaaac aatatctttt acatcattcg tatttaaaat tccaaactcc gctcccctaa
     1261 ggcgaataaa agccattaaa tcttttgtat ttaccaaatt atagtcatcc actatatcta
     1321 aaagtaaatt cttcaattct cttttttggc tttcatcaag tgttatatag cggtcaatat
     1381 caaaatcatt aatgttcaaa atatcttttt tgtcgtatat atgtttattc ttagcaatag
     1441 cgtcctttga ttcatgagtc aaatattcat atgaaccttt gatataatca agtatctcaa
     1501 catgagcaac tgaactattc cccaattttc gcttaatctt gttcctaacg ctttctattg
     1561 ttacaggatt tcgtgcaata tatataacgt gatagtgtgg ttttttatag tgctttccat
     1621 tttgtataat attactatta ttccatgtat ctttatcttt tttttcgtcc atatcgtgta
     1681 aaggactgac agccatagat acgcccaaac tctctaattt ttctttccaa tcattaggaa
     1741 ttgagtcagg atataataaa aatccaaaat ttctagcttt agtattttta atagccatga
     1801 tataattacc ttatcaaaaa caagtagcga aaactcgtat ccttctaaaa acgcgagctt
     1861 tcgcttattt tttttgttct gattcctttc ttgcatattc ttctatagct aacgccgcaa
     1921 ccgcagattt tgaaaaacct ttttgtttcg ccatatctgt taatttttta tcttgctctt
     1981 ttgtcagaga aatcataact ctttttttcg attctgaaat caccatttaa aaaactccaa
     2041 tcaaataatt ttataaagtt agtgtatcac tttgtaatca taaaaacaac aataaagcta
     2101 cttaaatata gatttataaa aaacgttggc gaaaacgttg gcgattcgtt ggcgattgaa
     2161 aaacccctta aacccttgag ccagttggga tagagcgttt ttggcacaaa aattggcact
     2221 cggcacttaa tggggggtcg tagtacggaa gcaaaattcg cttcctttcc ccccattttt
     2281 ttccaaattc caaatttttt tcaaaaattt tccagcgcta ccgctcggca aaattgcaag
     2341 caatttttaa aatcaaaccc atgagggaat ttcattccct catactccct tgagcctcct
     2401 ccaaccgaaa tagaagggcg ctgcgcttat tatttcattc agtcatcggc tttcataatc
     2461 taacagacaa catcttcgct gcaaagccac gctacgctca agggctttta cgctacgata
     2521 acgcctgttt taacgattat gccgataact aaacgaaata aacgctaaaa cgtctcagaa
     2581 acgcggccgc agctgttgac aattaatcat ccggctcgta taacgtgtgg aattgtgaag
     2641 aaaggaggtg aaatcatgag caatttgatt aacggaaaaa taccaaatca agcgattcaa
     2701 acattaaaaa tcgtaaaaga tttatttgga agttcaatag ttggagtata tctatttggt
     2761 tcagcagtaa atggtggttt acgcattaac agcgatgtag atgttctagt cgtcgtgaat
     2821 catagtttac ctcaattaac tcgaaaaaaa ctaacagaaa gactaatgac tatatcagga
     2881 aagattggaa atacggattc tgttagacca cttgaagtta cggttataaa taggagtgaa
     2941 gttgtccctt ggcaatatcc tccaaaaaga gaatttatat acggtgagtg gctcaggggt
     3001 gaatttgaga atggacaaat tcaggaacca agctatgatc ctgatttggc tattgtttta
     3061 gcacaagcaa gaaagaatag tatttctcta tttggtcctg attcttcaag tatacttgtc
     3121 tccgtacctt tgacagatat tcgaagagca attaaggatt ctttgccaga actaattgag
     3181 gggataaaag gtgatgagcg taatgtaatt ttaaccctag ctcgaatgtg gcaaacagtg
     3241 actactggtg aaattacctc gaaagatgtc gctgcagaat gggctatacc tcttttacct
     3301 aaagagcatg taactttact ggatatagct agaaaaggct atcggggaga gtgtgatgat
     3361 aagtgggaag gactatattc aaaggtgaaa gcactcgtta agtatatgaa aaattctata
     3421 gaaacttctc tcaattgatg gcaaaacaac ttgaaaaaag ttgttgacaa aaaagaagct
     3481 gaatgttata ttagtaaagt ctcgatgttt gttgacagta taaagttaga aacttataat
     3541 gataagaatt ttagaaagga ggtaatttag atgcgtggat caggatctat gagtgaattg
     3601 attaaagaaa atatgcatat gaaattatat atggaaggaa ctgttaataa tcatcatttc
     3661 aaatgtacat cagagggaga aggaaaacca tatgagggaa cgcaaactat gaaaataaag
     3721 gtggtagaag gagggccttt accatttgca ttcgatattt tagctacgag ttttatgtat
     3781 ggaagtaaag catttattaa tcatactcag gggataccgg atttctttaa acaatcattt
     3841 cctgaaggat ttacatggga acgtattaca acatatgaag atgggggagt attaacggca
     3901 actcaagata cgtcatttca aaatggatgc atcatataca atgttaaaat caatggggtt
     3961 aactttccat ctaatggacc tgtgatgcag aagaaaacac gtgggtggga agcgaatact
     4021 gagatgttat atccagctga tggagggtta agaggacata gtcaaatggc attgaaatta
     4081 gtaggagggg gatacttaca ttgttctttt aaaactacat atcgttctaa gaaaccagct
     4141 aaaaacttaa agatgccggg gtttcatttt gtagaccata gattagaacg tattaaagaa
     4201 gcggataaag aaacatatgt tgagcaacat gaaatggctg tagcaaaata ttgtgattta
     4261 cctagtaaat tagggcatag ataagaattc ccacgggttt tgctgcccgc aaacgggctg
     4321 ttctggtgtt gctagtttgt tatcagaatc gcagatccgg cttcgccagg tttgccggct
     4381 gaaagcgcta tttcttccag aattgccatg attttttccc cacgggaggc gtcactggct
     4441 cccgtgttgt cggcagcttt gattcgataa gcagcatcgc ctgtttcagg ctgtctatgt
     4501 gtgactgttg agctgtaaca agttgtctca ggtgttcaat ttcatgttct agttgctttg
     4561 ttttactggt ttcacctgtt ctattaggtg ttacatgctg ttcatctgtt acattgtcga
     4621 tctgttcatg gtgaacagct ttaaatgcac caaaaactcg taaaagctct gatgtatcta
     4681 tcttttttac accgttttca tctgtgcata tggacagttt tccctttgat atctaacggt
     4741 gaacagttgt tctacttttg tttgttagtc ttgatgcttc actgatagat acaagagcca
     4801 taagaacctc agatccttcc gtatttagcc agtatgttct ctagtgtggt tcgttgtttt
     4861 tgcgtgagcc atgagaacga accattgaga tcatgcttac tttgcatgtc actcaaaaat
     4921 tttgcctcaa aactggtgag ctgaattttt gcagttaaag catcgtgtag tgtttttctt
     4981 agtccgttac gtaggtagga atctgatgta atggttgttg gtattttgtc accattcatt
     5041 tttatctggt tgttctcaag ttcggttacg agatccattt gtctatctag ttcaacttgg
     5101 aaaatcaacg tatcagtcgg gcggcctcgc ttatcaacca ccaatttcat attgctgtaa
     5161 gtgtttaaat ctttacttat tggtttcaaa acccattggt taagcctttt aaactcatgg
     5221 tagttatttt caagcattaa catgaactta aattcatcaa ggctaatctc tatatttgcc
     5281 ttgtgagttt tcttttgtgt tagttctttt aataaccact cataaatcct catagagtat
     5341 ttgttttcaa aagacttaac atgttccaga ttatatttta tgaatttttt taactggaaa
     5401 agataaggca atatctcttc actaaaaact aattctaatt tttcgcttga gaacttggca
     5461 tagtttgtcc actggaaaat ctcaaagcct ttaaccaaag gattcctgat ttccacagtt
     5521 ctcgtcatca gctctctggt tgctttagct aatacaccat aagcattttc cctactgatg
     5581 ttcatcatct gagcgtattg gttataagtg aacgataccg tccgttcttt ccttgtaggg
     5641 ttttcaatcg tggggttgag tagtgccaca cagcataaaa ttagcttggt ttcatgctcc
     5701 gttaagtcat agcgactaat cgctagttca tttgctttga aaacaactaa ttcagacata
     5761 catctcaatt ggtctaggtg attttaatca ctataccaat tgagatgggc tagtcaatga
     5821 taattactag tccttttcct ttgagttgtg ggtatctgta aattctgcta gacctttgct
     5881 ggaaaacttg taaattctgc tagaccctct gtaaattccg ctagaccttt gtgtgttttt
     5941 tttgtttata ttcaagtggt tataatttat agaataaaga aagaataaaa aaagataaaa
     6001 agaatagatc ccagccctgt gtataactca ctactttagt cagttccgca gtattacaaa
     6061 aggatgtcgc aaacgctgtt tgctcctcta caaaacagac cttaaaaccc taaaggctta
     6121 agtagcaccc tcgcaagctc gggcaaatcg ctgaatattc cttttgtctc cgaccatcag
     6181 gcacctgagt cgctgtcttt ttcgtgacat tcagttcgct gcgctcacgg ctctggcagt
     6241 gaatgggggt aaatggcact acaggcgcct tttatggatt catgcaagga aactacccat
     6301 aatacaagaa aagcccgtca cgggcttctc agggcgtttt atggcgggtc tgctatgtgg
     6361 tgctatctga ctttttgctg ttcagcagtt cctgccctct gattttccag tctgaccact
     6421 tcggattatc ccgtgacagg tcattcagac tggctaatgc acccagtaag gcagcggtat
     6481 catcaacagg cttacccgtc ttactgtcgg gaattgatgc agaagcttga gctccta
//
