LOCUS       40924_20451        6872 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Temperature-sensitive shuttle vector pFNLTP9, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6872)
  AUTHORS   Maier TM, Havig A, Casey M, Nano FE, Frank DW, Zahrt TC.
  TITLE     Construction and characterization of a highly efficient Francisella 
            shuttle plasmid
  JOURNAL   Appl. Environ. Microbiol. 70 (12), 7511-7519 (2004)
  PUBMED    15574954
REFERENCE   2  (bases 1 to 6872)
  AUTHORS   Maier TM, Havig A, Casey M, Nano FE, Frank DW, Zahrt TC.
  TITLE     Direct Submission
  JOURNAL   Submitted (07-MAY-2004) Microbiology and Molecular Genetics, Medical
            College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI 
            53226, USA
REFERENCE   3  (bases 1 to 6872)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6872)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Appl. 
            Environ. Microbiol."; date: "2004"; volume: "70"; issue: "12"; 
            pages: "7511-7519"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (07-MAY-2004) Microbiology and Molecular Genetics, Medical College 
            of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI 53226, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6872
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        143..173
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    234..258
                     /note="multiple cloning site; MCS; derived from
                     pCR2.1-TOPO"
     mRNA            527..1420
                     /product="unknown"
                     /label=unknown mRNA
     CDS             542..1420
                     /codon_start=1
                     /product="unknown"
                     /label=unknown
                     /note="orf3"
                     /protein_id="AAT44340.1"
                     /translation="MKDKAAKNRDFRKTILSVLQRNKDGSFATQANRKSILLQATKDLK
                     KVGFSKVTAENFGNKHCYALRDHWRAQGLATATIKNRLACLRWLGEKMGKELPDNRKLE
                     IENRKYSDNSINKAQEIDFKAISALTDRQALAIQLQREFGLRREESLKFQPSYAIKEHK
                     IELKSSWTKGGRPREIPILNEKQRELLEKVKEVAGKGSLIESEKSYKQAMEHFTTRCQR
                     AGIKNVHGFRHAYAQDRYRQLTGRECPKNGGLTSKQLTPEQKQQDYEARMTISEELGHG
                     REDVTVNYLGR"
     CDS             complement(1637..2059)
                     /codon_start=1
                     /product="RepB protein"
                     /label=RepB protein
                     /note="orf2"
                     /protein_id="AAT44341.1"
                     /translation="MNIEIFKFSKKHKKINKKLIDLTKVKQVVTTGYSFIVVYQRGRKI
                     ELSYESSYWHAKKDLDNSNSDVGITYSYIKKDKRGPCHKDNYLRDSKGRLLKDWGKIEE
                     LHEKYPKIGKIWYISREETDKLLKDFTYKNGNLIMV"
     gene            complement(2062..3139)
                     /gene="repA"
                     /label=repA
     mRNA            complement(2062..3104)
                     /gene="repA"
                     /product="mutant RepA protein"
                     /label=repA mRNA
     CDS             complement(2062..3081)
                     /codon_start=1
                     /gene="repA"
                     /product="mutant RepA protein"
                     /label=repA
                     /note="orf1; RepA M120I"
                     /protein_id="AAT44342.1"
                     /translation="MKEVKNVNAGKEIAMSNTLVAGKYSLTKEEQNLIFLVASMVKRED
                     KEFHRYKISLSDLEKATGVKHNRVRLKQLMHSIMSKPVWLNKEQTKIANWFAYIEADPK
                     SSALICEFHWSLMPHIIQLQEYFTKAERQLLFSFKSKYSSRLYLLLKSKLGEQAGYANI
                     VDCVLYVDDMINDFDLPKSYSNRYSNFKNKFLLPALEEINQLSDIFVTYDDKDHHRKHG
                     RKITNIKFTVSKVAETEEEFKQKLLDTKNKEDYIPIEASERLREVVLSDELDLDLYYVR
                     SIFRHHHLHDIERVCDDTWRNWDNKKIKDRKRVFIANIKKLEKKPEENHKLPFGFEEV"
     variation       2722
                     /gene="repA"
                     /replace="c"
                     /note="compared to pFNLTP1 wild-type repA in GenBank
                     Accession Number AY622904; results in isoleucine to 
                     methionine amino acid change"
     regulatory      complement(3111..3116)
                     /gene="repA"
                     /regulatory_class="minus_10_signal"
     regulatory      complement(3134..3139)
                     /gene="repA"
                     /regulatory_class="minus_35_signal"
     repeat_region   3168..3262
     repeat_region   3317..3347
     CDS             3452..3637
                     /codon_start=1
                     /product="unknown"
                     /label=unknown
                     /note="orf5; hybrid ORF; contains partial phd sequence"
                     /protein_id="AAT44343.1"
                     /translation="MQTVNYSTFRNELSDSMDRVTKNHSPMIVTRGSKKEAVVMMSLED
                     SSLPFSPRSPAFPVKL"
     CDS             4260..5051
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
     CDS             5072..5929
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      6103..6691
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcttg
      241 gtaccgagct cggatccact agctcgtttc aaattaccga tgatatcgga ccgttccaac
      301 ttaccgacca gttcggcagg tatgtatttg cgtgcattcc tatccaaaaa aacatcaagc
      361 caaaagcttg aaaaaactta caacacagct caacagagct agattgtaaa accctgcttt
      421 gttaagcaga acgcaaaaat tgaatgactt atagtcatat cgcttcgacc ctcgtagatt
      481 agtagccttg agctattaac tggttgaaac acttaccaaa taaagattaa aagcgataaa
      541 aatgaaagat aaagcagcta aaaacagaga ttttagaaag actattttat cagtgttaca
      601 acgcaataaa gatggctctt ttgctacgca agcaaataga aagtctattc tgttgcaggc
      661 aactaaagac cttaaaaagg tagggtttag caaggttaca gccgaaaact tcggtaataa
      721 gcattgctat gcacttagag accattggag agcccaagga ttagctacag caacgataaa
      781 aaatcgttta gcttgtctaa ggtggttagg cgagaaaatg ggcaaagaac tacccgataa
      841 tcgaaaatta gagattgaga acaggaagta tagcgataat tcaatcaata aagcccaaga
      901 aatcgatttt aaggcgattt ctgccttaac tgataggcaa gccctagcaa tacaattaca
      961 gcgcgaattt gggcttcgta gagaagaaag tttgaagttt cagcctagtt atgcaatcaa
     1021 agagcataaa atcgagctta aaagctcttg gacaaagggt ggaagaccac gagaaatccc
     1081 aattttgaat gaaaaacaga gagaattgtt agaaaaagta aaagaggtag caggtaaagg
     1141 ctctctaatt gagagcgaaa agtcttataa gcaagcaatg gaacatttca cgactcgctg
     1201 tcaaagagca gggattaaga atgttcatgg ctttagacat gcgtatgctc aagatagata
     1261 taggcaatta acagggcgtg agtgtcctaa aaatggtgga ttaacatcta agcagctaac
     1321 acctgagcaa aagcaacaag actatgaagc tagaatgact attagtgaag agttaggtca
     1381 tggtagagaa gatgtaacag tcaactactt aggcagataa aaagcaatat agctatagaa
     1441 gaaaagaaag ctattttaca tagtagatcg actcttctta gggattttat attttttgat
     1501 aaatcatcta ttttgctagt taaatcatca aatttatcat cttgttgttt gactaaatct
     1561 aagaatctat tctctttttt aaaatcgttc atgcaaaccg cctatagctt tcttcttttt
     1621 ctgaaattat ttgtcttcac accataatta aattcccatt tttataagta aagtctttta
     1681 aaagcttgtc agtctcttct ctagaaatgt accaaatttt acctatttta ggatactttt
     1741 catgaagctc ttctattttt ccccagtcct ttaatagtct acctttagag tctcgtaaat
     1801 agttatcttt gtgacagggg cctcttttat cttttttaat gtaactatat gttattccaa
     1861 cgtcactatt actattatcc aaatcttttt tagcatgcca gtaagaactt tcataactta
     1921 actctatctt tcgacctctt tgatatacaa caataaagct atagccagta gtaacaacct
     1981 gttttacttt tgttaaatct attaacttct tatttatttt tttatgtttt tttgaaaatt
     2041 taaatatttc tatattcatt cctacacttc ctcaaatcca aatggtagct tatgattctc
     2101 ttctggtttc ttttctaatt tttttatatt tgcaataaaa actctttttc tatctttgat
     2161 ttttttattg tcccaattcc tccaagtatc atcacaaacc ctttcaatat catgtaaatg
     2221 atgatgtcta aatattgatc tgacataata cagatctagg tctagttcat cacttaacac
     2281 aacttctcta agtctttcag atgcttcgat tggtatgtaa tcctctttat ttttagtatc
     2341 taaaagcttt tgcttaaatt cttcttctgt ctctgctacc ttactaactg taaacttgat
     2401 atttgtaatc ttacgaccat gttttctgtg atgatccttg tcatcatagg ttacaaaaat
     2461 atccgataat tgattaatct cttctagtgc tggtaatagg aacttatttt taaaatttga
     2521 atatctgttg ctgtaacttt taggtaaatc aaaatcatta atcatatcat cgacatacaa
     2581 tacgcaatca actatattag cataccctgc ttgttcgcct aatttgcttt tgagaagtaa
     2641 gtataatctg cttgaatact tacttttaaa tgaaaatagt aactgtcttt ctgctttagt
     2701 aaagtactct tgtagttgta ttatgtgtgg cattaatgac caatgaaact cgcaaattaa
     2761 agcactgctt ttagggtctg cttcaatata tgcaaaccag ttagctatct tcgtttgttc
     2821 tttattcagc catactggct tagacattat tgagtgcatt aattgcttca atctcactct
     2881 gttatgctta acccctgtag ctttttcaag atcagatagg cttatcttat acctgtgaaa
     2941 ctctttatct tctcttttaa ccattgaggc aactaagaat attaagtttt gttcttcttt
     3001 tgtaaggcta tactttcctg caacaagagt attagacata gctatttctt tgccagcatt
     3061 tacattttta acttctttca tagaactaga gtcattatct cgatatacaa attctataaa
     3121 acttctatta gtaaaacaac tacttcataa aaaaaagtag ttttaacgat acaaaaagta
     3181 gttttaaatt caaaaagtga tacaaaaagt agttttaaat tcaaaaagtg atacaaaaag
     3241 tagttttaaa ttttttaaaa aagtgcttca aagccttatg tagcaatact tacagaggat
     3301 taaaaaaaaa tctgacaata tataaagaga atatataaag agaatatctt aggggatttt
     3361 aaaaaaatcc cacagactca aagacttttt tgacttttta aatcctagaa actatacttt
     3421 aagtacttat ttaagtacat ggatttagat tatgcaaacc gttaattatt caacttttag
     3481 aaatgaacta tctgattcaa tggatagagt aacaaaaaat catagtccta tgattgtaac
     3541 tagaggttca aaaaaagaag cagttgttat gatgtcgtta gaggattctt cccttccttt
     3601 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     3661 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     3721 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     3781 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     3841 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     3901 aaatttaacg cgaattttaa caaaattcag ggcgcaaggg ctgctaaagg aagcggaaca
     3961 cgtagaaagc cagtccgcag aaacggtgct gaccccggat gaatgtcagc tactgggcta
     4021 tctggacaag ggaaaacgca agcgcaaaga gaaagcaggt agcttgcagt gggcttacat
     4081 ggcgatagct agactgggcg gttttatgga cagcaagcga accggaattg ccagctgggg
     4141 cgccctctgg taaggttggg aagccctgca aagtaaactg gatggctttc ttgccgccaa
     4201 ggatctgatg gcgcagggga tcaagatctg atcaagagac aggatgagga tcgtttcgca
     4261 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     4321 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggttgtcag
     4381 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     4441 aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     4501 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     4561 atctcctgtc atcccacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     4621 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     4681 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     4741 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg
     4801 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     4861 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     4921 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     4981 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     5041 acgagttctt ctgaattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     5101 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     5161 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     5221 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     5281 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     5341 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     5401 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     5461 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     5521 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     5581 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     5641 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     5701 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     5761 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     5821 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     5881 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     5941 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     6001 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     6061 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     6121 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     6181 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     6241 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     6301 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     6361 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     6421 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     6481 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     6541 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     6601 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     6661 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     6721 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     6781 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     6841 gcgcagcgag tcagtgagcg aggaagcgga ag
//
