LOCUS       V004420                 9196 bp    DNA     circular SYN 18-DEC-2018
DEFINITION  Exported.
ACCESSION   V004420
VERSION     V004420
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 9196)
  AUTHORS   Heap JT, Kuehne SA, Ehsaan M, Cartman ST, Cooksley CM, Scott JC,
            Minton NP.
  TITLE     The ClosTron: Mutagenesis in Clostridium refined and streamlined
  JOURNAL   J. Microbiol. Methods 80 (1), 49-55 (2010)
   PUBMED   19891996
REFERENCE   2  (bases 1 to 9196)
  AUTHORS   Heap JT, Kuehne KA, Ehsaan M, Cartman ST, Cooksley CM, Minton NP.
  TITLE     Direct Submission
  JOURNAL   Submitted (28-SEP-2010) School of Molecular Medical Sciences, The
            University of Nottingham, Centre for Biomolecular Sciences,
            Nottingham, Nottinghamshire NG72RD, UK
REFERENCE   3  (bases 1 to 9196)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 9196)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J.
            Microbiol. Methods"; date: "2010"; volume: "80"; issue: "1"; pages:
            "49-55"
            SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
            (28-SEP-2010) School of Molecular Medical Sciences, The University
            of Nottingham, Centre for Biomolecular Sciences, Nottingham,
            Nottinghamshire NG72RD, UK"
            SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..9196
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             233..406
                     /label="lacZ-alpha"
                     /note="LacZ-alpha fragment of beta-galactosidase"
     protein_bind    complement(814..847)
                     /label="FRT (minimal)"
                     /bound_moiety="FLP recombinase from the Saccharomyces
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration
                     (Turan and Bode, 2011)"
     CDS             complement(864..1598)
                     /gene="ermBP"
                     /label="rRNA adenine N-6-methyltransferase"
                     /note="rRNA adenine N-6-methyltransferase from Enterococcus
                     faecalis. Accession#: P0A4D5"
     protein_bind    complement(2101..2134)
                     /label="FRT (minimal)"
                     /note="supports FLP-mediated excision but not integration
                     (Turan and Bode, 2011)"
     RBS             2392..2414
                     /label="RBS"
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2606..4402
                     /gene="ltrA"
                     /label="Group II intron-encoded protein LtrA"
                     /note="Group II intron-encoded protein LtrA from
                     Lactococcus lactis subsp. cremoris (strain MG1363).
                     Accession#: P0A3U1"
     terminator      4590..4676
                     /label="rrnB T1 terminator"
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      4768..4795
                     /label="rrnB T2 terminator"
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     rep_origin      7640..8228
                     /direction=RIGHT
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     oriT            8549..8658
                     /label="oriT"
                     /note="incP origin of transfer"
     CDS             8691..9059
                     /label="traJ"
                     /note="oriT-recognizing protein"
ORIGIN
        1 cctgcagggt gtagtagcct gtgaaataag taaggaaaaa aaagaagtaa gtgttatata
       61 tgatgattat tttgtagatg tagataggat aatagaatcc atagaaaata taggttatac
      121 agttatataa aaattacttt aaaaattaat aaaaacatgg taaaatataa atcgtataaa
      181 gttgtgtaat ttttaagctt gagctcataa caatttcaca caggaaacag ctatgaccat
      241 gattacggat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
      301 ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc
      361 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc gctaataaag
      421 atcttgtaca atctgtagga gaacctatgg gaacgaaacg aaagcgatgc cgagaatctg
      481 aatttaccaa gacttaacac taactgggga taccctaaac aagaatgcct aatagaaagg
      541 aggaaaaagg ctatagcact agagcttgaa aatcttgcaa gggtacggag tactcgtagt
      601 agtctgagaa gggtaacgcc ctttacatgg caaaggggta cagttattgt gtactaaaat
      661 taaaaattga ttagggagga aaacctcaaa atgaaaccaa caatggcaat tttagaaaga
      721 atcagtaaaa attcacaaga aaatatagac gaagttttta caagacttta tcgttatctt
      781 ttacgtccag atatttatta cgtggcgacg cgtgaagttc ctatactttc tagagaatag
      841 gaacttcgcg actcatagaa ttatttcctc ccgttaaata atagataact attaaaaata
      901 gacaatactt gctcataagt aacggtactt aaattgttta ctttggcgtg tttcattgct
      961 tgatgaaact gatttttagt aaacagttga cgatattctc gattgaccca ttttgaaaca
     1021 aagtacgtat atagcttcca atatttatct ggaacatctg tggtatggcg ggtaagtttt
     1081 attaagacac tgtttacttt tggtttagga tgaaagcatt ccgctggcag cttaagcaat
     1141 tgctgaatcg agacttgagt gtgcaagagc aaccctagtg ttcggtgaat atccaaggta
     1201 cgcttgtaga atccttcttc aacaatcaga tagatgtcag acgcatggct ttcaaaaacc
     1261 acttttttaa taatttgtgt gcttaaatgg taaggaatac tcccaacaat tttatacctc
     1321 tgtttgttag ggaattgaaa ctgtagaata tcttggtgaa ttaaagtgac acgagtattc
     1381 agttttaatt tttctgacga taagttgaat agatgactgt ctaattcaat agacgttacc
     1441 tgtttactta ttttagccag tttcgtcgtt aaatgccctt tacctgttcc aatttcgtaa
     1501 acggtatcgg tttcttttaa attcaattgt tttattattt ggttgagtac tttttcactc
     1561 gttaaaaagt tttgagaata ttttatattt ttgttcatac cagcaccaga agcaccagca
     1621 tctcttgggt taattgaggc ctgagtataa ggtgacttat acttgtaatc tatctaaacg
     1681 gggaacctct ctagtagaca atcccgtgct aaattgtagg actgcccttt aataaatact
     1741 tctatattta aagaggtatt tatgaaaagc ggaatttatc agattaaaaa tactttctct
     1801 agagaaaatt tcgtctggat tagttactta tcgtgtaaaa tctgataaat ggaattggtt
     1861 ctacataaat gcctaacgac tatccctttg gggagtaggg tcaagtgact cgaaacgata
     1921 gacaacttgc tttaacaagt tggagatata gtctgctctg catggtgaca tgcagctgga
     1981 tataattccg gggtaagatt aacgacctta tctgaacata atgccatatg aatccctcct
     2041 aatttatacg ttttctctaa caacttaatt atacccacta ttattatttt tatcaatata
     2101 gaagttccta tactttctag agaataggaa cttcacgcgt tgggaaatgg caatgatagc
     2161 gaaacaacgt aaaactcttg ttgtatgctt tcattgtcat cgtcacgtga ttcataaaca
     2221 caagtgaatg tcgacagtga atttttacga acgaacaata acagagccgt atactccgag
     2281 aggggtacgt acggttcccg aagagggtgg tgcaaaccag tcacagtaat gtgaacaagg
     2341 cggtacctcc ctacttcacc atatcatttt ctgcagcccc ctagaaataa ttttgtttaa
     2401 ctttaagaag gagatataca tatatggcta gatcgtccat tccgacagca tcgccagtca
     2461 ctatggcgtg ctgctagcgc tatatgcgtt gatgcaattt ctatgcactc gtagtagtct
     2521 gagaagggta acgcccttta catggcaaag gggtacagtt attgtgtact aaaattaaaa
     2581 attgattagg gaggaaaacc tcaaaatgaa accaacaatg gcaattttag aaagaatcag
     2641 taaaaattca caagaaaata tagacgaagt ttttacaaga ctttatcgtt atcttttacg
     2701 tccagatatt tattacgtgg cgtatcaaaa tttatattcc aataaaggag cttccacaaa
     2761 aggaatatta gatgatacag cggatggctt tagtgaagaa aaaataaaaa agattattca
     2821 atctttaaaa gacggaactt actatcctca acctgtacga agaatgtata ttgcaaaaaa
     2881 gaattctaaa aagatgagac ctttaggaat tccaactttc acagataaat tgatccaaga
     2941 agctgtgaga ataattcttg aatctatcta tgaaccggta ttcgaagatg tgtctcacgg
     3001 ttttagacct caacgaagct gtcacacagc tttgaaaaca atcaaaagag agtttggcgg
     3061 cgcaagatgg tttgtggagg gagatataaa aggctgcttc gataatatag accacgttac
     3121 actcattgga ctcatcaatc ttaaaatcaa agatatgaaa atgagccaat tgatttataa
     3181 atttctaaaa gcaggttatc tggaaaactg gcagtatcac aaaacttaca gcggaacacc
     3241 tcaaggtgga attctatctc ctcttttggc caacatctat cttcatgaat tggataagtt
     3301 tgttttacaa ctcaaaatga agtttgaccg agaaagtcca gaaagaataa cacctgaata
     3361 tcgggagctc cacaatgaga taaaaagaat ttctcaccgt ctcaagaagt tggagggtga
     3421 agaaaaagct aaagttcttt tagaatatca agaaaaacgt aaaagattac ccacactccc
     3481 ctgtacctca cagacaaata aagtattgaa atacgtccgg tatgcggacg acttcattat
     3541 ctctgttaaa ggaagcaaag aggactgtca atggataaaa gaacaattaa aactttttat
     3601 tcataacaag ctaaaaatgg aattgagtga agaaaaaaca ctcatcacac atagcagtca
     3661 acccgctcgt tttctgggat atgatatacg agtaaggaga tctggaacga taaaacgatc
     3721 tggtaaagtc aaaaagagaa cactcaatgg gagtgtagaa ctccttattc ctcttcaaga
     3781 caaaattcgt caatttattt ttgacaagaa aatagctatc caaaagaaag atagctcatg
     3841 gtttccagtt cacaggaaat atcttattcg ttcaacagac ttagaaatca tcacaattta
     3901 taattctgaa ctccgcggga tttgtaatta ctacggtcta gcaagtaatt ttaaccagct
     3961 caattatttt gcttatctta tggaatacag ctgtctaaaa acgatagcct ccaaacataa
     4021 gggaacactt tcaaaaacca tttccatgtt taaagatgga agtggttcgt gggggatccc
     4081 gtatgagata aagcaaggta agcagcgccg ttattttgca aattttagtg aatgtaaatc
     4141 cccttatcaa tttacggatg agataagtca agctcctgta ttgtatggct atgcccggaa
     4201 tactcttgaa aacaggttaa aagctaaatg ttgtgaatta tgtgggacgt ctgatgaaaa
     4261 tacttcctat gaaattcacc atgtcaataa ggtcaaaaat cttaaaggca aagaaaaatg
     4321 ggaaatggca atgatagcga aacaacgtaa aactcttgtt gtatgctttc attgtcatcg
     4381 tcacgtgatt cataaacaca agtgaatgtc gagcacccgt tctcggagca ctgtccgacc
     4441 gctttggccg ccgcccagtc ctgctcgctt cgctacttgg agccactatc gactacgcga
     4501 tcatggcgac cacacccgtc ctgtggatcg ccaagccgcc gatggtagtg tggggtctcc
     4561 ccatgcgaga gtagggaact gccaggcatc aaataaaacg aaaggctcag tcgaaagact
     4621 gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc
     4681 cgggagcgga tttgaacgtt gcgaagcaac ggcccggagg gtggcgggca ggacgcccgc
     4741 cataaactgc caggcatcaa attaagcaga aggccatcct gacggatggc ctttttgcgt
     4801 ttctacaaac tcttcctgtc gtcatatcta caagccatcc ccccacagat acgggcgcgc
     4861 cgccattatt tttttgaaca attgacaatt catttcttat tttttattaa gtgatagtca
     4921 aaaggcataa cagtgctgaa tagaaagaaa tttacagaaa agaaaattat agaatttagt
     4981 atgattaatt atactcattt atgaatgttt aattgaatac aaaaaaaaat acttgttatg
     5041 tattcaatta cgggttaaaa tatagacaag ttgaaaaatt taataaaaaa ataagtcctc
     5101 agctcttata tattaagcta ccaacttagt atataagcca aaacttaaat gtgctaccaa
     5161 cacatcaagc cgttagagaa ctctatctat agcaatattt caaatgtacc gacatacaag
     5221 agaaacatta actatatata ttcaatttat gagattatct taacagatat aaatgtaaat
     5281 tgcaataagt aagatttaga agtttatagc ctttgtgtat tggaagcagt acgcaaaggc
     5341 ttttttattt gataaaaatt agaagtatat ttattttttc ataattaatt tatgaaaatg
     5401 aaagggggtg agcaaagtga cagaggaaag cagtatctta tcaaataaca aggtattagc
     5461 aatatcatta ttgactttag cagtaaacat tatgactttt atagtgcttg tagctaagta
     5521 gtacgaaagg gggagcttta aaaagctcct tggaatacat agaattcata aattaattta
     5581 tgaaaagaag ggcgtatatg aaaacttgta aaaattgcaa agagtttatt aaagatactg
     5641 aaatatgcaa aatacattcg ttgatgattc atgataaaac agtagcaacc tattgcagta
     5701 aatacaatga gtcaagatgt ttacataaag ggaaagtcca atgtattaat tgttcaaaga
     5761 tgaaccgata tggatggtgt gccataaaaa tgagatgttt tacagaggaa gaacagaaaa
     5821 aagaacgtac atgcattaaa tattatgcaa ggagctttaa aaaagctcat gtaaagaaga
     5881 gtaaaaagaa aaaataattt atttattaat ttaatattga gagtgccgac acagtatgca
     5941 ctaaaaaata tatctgtggt gtagtgagcc gatacaaaag gatagtcact cgcattttca
     6001 taatacatct tatgttatga ttatgtgtcg gtgggacttc acgacgaaaa cccacaataa
     6061 aaaaagagtt cggggtaggg ttaagcatag ttgaggcaac taaacaatca agctaggata
     6121 tgcagtagca gaccgtaagg tcgttgttta ggtgtgttgt aatacatacg ctattaagat
     6181 gtaaaaatac ggataccaat gaagggaaaa gtataatttt tggatgtagt ttgtttgttc
     6241 atctatgggc aaactacgtc caaagccgtt tccaaatctg ctaaaaagta tatcctttct
     6301 aaaatcaaag tcaagtatga aatcataaat aaagtttaat tttgaagtta ttatgatatt
     6361 atgtttttct attaaaataa attaagtata tagaatagtt taataatagt atatacttaa
     6421 tgtgataagt gtctgacagt gtcacagaaa ggatgattgt tatggattat aagcggccgg
     6481 cccaatgaat aggtttacac ttactttagt tttatggaaa tgaaagatca tatcatatat
     6541 aatctagaat aaaattaact aaaataatta ttatctagat aaaaaattta gaagccaatg
     6601 aaatctataa ataaactaaa ttaagtttat ttaattaaca actatggata taaaataggt
     6661 actaatcaaa atagtgagga ggatatattt gaatacatac gaacaaatta ataaagtgaa
     6721 aaaaatactt cggaaacatt taaaaaataa ccttattggt acttacatgt ttggatcagg
     6781 agttgagagt ggactaaaac caaatagtga tcttgacttt ttagtcgtcg tatctgaacc
     6841 attgacagat caaagtaaag aaatacttat acaaaaaatt agacctattt caaagaaaat
     6901 aggagataaa agcaacttac gatatattga attaacaatt attattcagc aagaaatggt
     6961 accgtggaat catcctccca aacaagaatt tatttatgga gaatggttac aagagcttta
     7021 tgaacaagga tacattcctc agaaggaatt aaattcagat ttaaccataa tgctttacca
     7081 agcaaaacga aaaaataaaa gaatatacgg aaattatgac ttagaggaat tactacctga
     7141 tattccattt tctgatgtga gaagagccat tatggattcg tcagaggaat taatagataa
     7201 ttatcaggat gatgaaacca actctatatt aactttatgc cgtatgattt taactatgga
     7261 cacgggtaaa atcataccaa aagatattgc gggaaatgca gtggctgaat cttctccatt
     7321 agaacatagg gagagaattt tgttagcagt tcgtagttat cttggagaga atattgaatg
     7381 gactaatgaa aatgtaaatt taactataaa ctatttaaat aacagattaa aaaaattata
     7441 aaaaaattga aaaaatggtg gaaacacttt tttcaatttt tttgttttat tatttaatat
     7501 ttgggaaata ttcattctaa ttggtaatca gattttagaa gtttaaactc ctttttgata
     7561 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     7621 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     7681 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     7741 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     7801 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     7861 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     7921 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     7981 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     8041 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     8101 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     8161 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     8221 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     8281 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     8341 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     8401 aagcggaaga gcgcccaata cgcagggccc cctgcttcgg ggtcattata gcgatttttt
     8461 cggtatatcc atcctttttc gcacgatata caggattttg ccaaagggtt cgtgtagact
     8521 ttccttggtg tatccaacgg cgtcagccgg gcaggatagg tgaagtaggc ccacccgcga
     8581 gcgggtgttc cttcttcact gtcccttatt cgcacctggc ggtgctcaac gggaatcctg
     8641 ctctgcgagg ctggccggct accgccggcg taacagatga gggcaagcgg atggctgatg
     8701 aaaccaagcc aaccaggaag ggcagcccac ctatcaaggt gtactgcctt ccagacgaac
     8761 gaagagcgat tgaggaaaag gcggcggcgg ccggcatgag cctgtcggcc tacctgctgg
     8821 ccgtcggcca gggctacaaa atcacgggcg tcgtggacta tgagcacgtc cgcgagctgg
     8881 cccgcatcaa tggcgacctg ggccgcctgg gcggcctgct gaaactctgg ctcaccgacg
     8941 acccgcgcac ggcgcggttc ggtgatgcca cgatcctcgc cctgctggcg aagatcgaag
     9001 agaagcagga cgagcttggc aaggtcatga tgggcgtggt ccgcccgagg gcagagccat
     9061 gactttttta gccgctaaaa cggccggggg gtgcgcgtga ttgccaagca cgtccccatg
     9121 cgctccatca agaagagcga cttcgcggag ctggtgaagt acatcaccga cgagcaaggc
     9181 aagaccgatc gggccc
//
