LOCUS       40924_33937        7395 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Chromosomal integration vector pOSIP-TT, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7395)
  AUTHORS   St-Pierre F, Cui L, Priest DG, Endy D, Dodd IB, Shearwin KE.
  TITLE     One-step cloning and chromosomal integration of DNA
  JOURNAL   ACS Synth Biol 2 (9), 537-541 (2013)
  PUBMED    24050148
REFERENCE   2  (bases 1 to 7395)
  AUTHORS   St-Pierre F, Cui L, Priest DG, Endy D, Dodd IB, Shearwin KE.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-MAY-2013) Molecular and Biomedical Science, The 
            University of Adelaide, Molecular Life Sciences Building, Adelaide, 
            South Australia 5005, Australia
REFERENCE   3  (bases 1 to 7395)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7395)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "ACS Synth 
            Biol"; date: "2013"; volume: "2"; issue: "9"; pages: "537-541"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (12-MAY-2013) Molecular and Biomedical Science, The University of 
            Adelaide, Molecular Life Sciences Building, Adelaide, South 
            Australia 5005, Australia"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7395
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             142..444
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
     rep_origin      477..1065
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    1125..1145
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1200..1227
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(1276..1307)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription
                     terminator"
     protein_bind    1384..1417
                     /label=FRT (minimal)
                     /note="supports FLP-mediated excision but not integration
                     (Turan and Bode, 2011)"
     CDS             complement(1427..2137)
                     /label=lambda repressor (ts)
                     /note="temperature-sensitive variant of the phage lambda
                     repressor"
     RBS             2226..2234
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             2299..3438
                     /label=phage phi-21 integrase
                     /note="integrase from phage phi-21"
     protein_bind    3622..3640
                     /label=tet operator
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             3649..4851
                     /label=TcR
                     /note="tetracycline efflux protein"
     terminator      4913..5007
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      5064..5452
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K;
                     requires the R6K initiator protein pi for replication"
     protein_bind    5463..5510
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     misc_feature    5477..5510
                     /note="FRT; Flp recombinase target site"
     protein_bind    5590..6089
                     /label=phage phi-21 attP
                     /note="attachment site of phage phi-21"
     terminator      complement(6110..6356)
                     /label=lambda tL3 terminator
                     /note="transcription terminator tL3 from phage lambda"
     terminator      6676..6762
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      6857..6943
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      7038..7124
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      complement(7222..7265)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
ORIGIN
        1 gaattcgagc tcggtacccg gggatcctct agagggctta ctaaaagcca gataacagta
       61 tgcgtatttg cgcgctgatt tttgcggtat aagaatatat actgatatgt atacccgaag
      121 tatgtcaaaa agaggtatgc tatgcagttt aaggtttaca cctataaaag agagagccgt
      181 tatcgtctgt ttgtggatgt acagagtgat attattgaca cgcccgggcg acggatggtg
      241 atccccctgg ccagtgcacg tctgctgtca gataaagtct cccgtgaact ttacccggtg
      301 gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg atatggccag tgtgccggtc
      361 tccgttatcg gggaagaagt ggctgatctc agccaccgcg aaaatgacat caaaaacgcc
      421 attaacctga tgttctgggg aatataataa cccgtagaaa agatcaaaag atcttcttga
      481 gatccttttt ttctgcgcgt aatctgctac ttgcaaacaa aaaaaccacc gctaccagcg
      541 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggctttagc
      601 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
      661 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
      721 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
      781 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
      841 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
      901 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
      961 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     1021 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     1081 gcctttttac ggttcctggc cttttgctgg ccttttgctc acattactag tagcggccgc
     1141 tgcaggcatg cctcgagatg catggcgcct aacctaaact gacaggcatc aaattaagca
     1201 gaaggccatc ctgacggatg gcctttttgc gtttcgaaca attgaaaaaa cctcgcgcct
     1261 tacctgttga gtaatagtca aaagcctccg gtcggaggct tttgactttc tgcttactga
     1321 atttcggtgg tgccgttaat taaccggtgg gccctcatga taataatggt ttcttagacg
     1381 tccgaagttc ctattctcta gaaagtatag gaacttccct aggtcagcca aacgtctctt
     1441 caggccactg actagcgata actttcccca caacggaaca actctcattg catgggatca
     1501 ttgggtactg tgggtttagt ggttgtaaaa acacctgacc gctatccctg atcagtttct
     1561 tgaaggtaaa ctcatcaccc ccaagtctgg ctatgcagaa atcacctggc tcaacagcct
     1621 gctcagggtc aacgagaatt aacattccgt caggaaagct tggcttggag cctgttggtg
     1681 cggtcatgga attaccttca acctcaagcc agaatgcaga atcactggct tttttggttg
     1741 tgcttaccca tctctccgca tcacctttgg taaaggttct aagcttaggt gagaacatcc
     1801 ctgcctgaac atgagaaaaa acagggtact catactcact tctaagtgac ggctgcatac
     1861 taaccgcttc atacatctcg tagatttctc tggcgattga agggctaaat tcttcaacgc
     1921 taactttgag aatttttgta agcaatgcgg cgttataagc atttaatgca ttgatgccat
     1981 taaataaagc accaacgcct gactgcccca tccccatctt gtctgcgaca gattcctggg
     2041 ataagccaag ttcatttttc tttttttcat aaattgcttt aaggcgacgt gcgtcctcaa
     2101 gctgctcttg tgttaatggt ttcttttttg tgctcatacg ttaaatctat caccgcaagg
     2161 gataaatatc taacaccgtg cgtgttgact attttacctc tggcggtgat aatggttgca
     2221 tgtactaagg aggttgtata cgcgtagaac cgcaactccc aataaacgca aacccaaaac
     2281 tccaacggat aatcgctgat ggctgctaga ccccgatctc acaaaatctc tatacccaat
     2341 ttatattgca aattagataa gcgaaccgga aaggtatatt ggcaatacaa acatccacta
     2401 tccggtcgtt ttcatagctt aggaactgat gagaatgaag caaaacaagt tgctactgaa
     2461 gcaaatacca ttattgctga acaacgtacc agacaaatat taagcgtcaa tgagcgtctg
     2521 gaaagaatga aaggcaggcg ctcagacatt acggtgacag aatggcttga taaatatatt
     2581 tctatccagg aggacaggct gcaacataat gaactaagac ccaactccta tcggcaaaaa
     2641 ggcaaaccca ttcgtctttt ccgtgagcat tgtggaatgc aacacctcaa ggatattacc
     2701 gcacttgata ttgccgaaat aattgatgct gtaaaggctg aaggtcataa caggatggcg
     2761 caagtcgtga gaatggtgtt gatcgacgtc ttcaaagaag cacaacacgc aggacatgtt
     2821 ccgccaggat ttaacccagc gcaggcaaca aaacaaccgc gaaatcgagt aaaccgccaa
     2881 agattgtcac tgctcgaatg gcaggcaata tttgaaagcg taagcagacg gcagccctat
     2941 ttaaaatgcg gcatgctact tgctcttgtt actggacaac gtttaggcga tatctgcaat
     3001 ttgaaattct ctgatatatg ggacgacatg ttgcacatta ctcaggaaaa aaccggttca
     3061 aaacttgcta ttccgcttaa cctgaaatgc gatgctctga atattaccct tcgtgaagtt
     3121 atatctcagt gcagggatgc tgttgttagt aaatatctgg tccattaccg tcacactacc
     3181 tctcaagcaa acagaggaga ccaggtttct gcaaatactc tgacaacggc ttttaaaaag
     3241 gccaaggaaa aatgtggcat aaaatgggag caaggaactg cgcccacatt tcatgaacag
     3301 cgatctctgt cagaacgatt atatcgggaa cagggtctgg atacgcaaaa gttgttaggc
     3361 cataaatcca gaaaaatgac cgaccgatac aatgatgatc gtggtaaaga ctggattatc
     3421 gtagatatca aaacagcata gaaaatagcc agttttgggg aagggttttg gggaaagttt
     3481 tggggaagat tttacatcat cataaaacaa cgggcgtgtt atacgcccgt tatccatggg
     3541 tatggacagt tttccctttg atatgtaacg cacgttgtgc atcgatttga caccctatca
     3601 ttgatagagt tattttccca ctccctatca gtgatagaga aaagtgaaat gaatagttcg
     3661 acaaagatcg cattggtaat tacgttactc gatgccatgg ggattggcct tatcatgcca
     3721 gtcttgccaa cgttattacg tgaatttatt gcttcggaag atatcgctaa ccactttggc
     3781 gtattgcttg cactttatgc gttaatgcag gttatctttg ctccttggct tggaaaaatg
     3841 tctgaccgat ttggtcggcg cccagtgctg ttgttgtcat taataggcgc atcgctggat
     3901 tacttattgc tggctttttc aagtgcgctt tggatgctgt atttaggccg tttgctttca
     3961 gggatcacag gagctactgg ggctgtcgcg gcatcggtca ttgccgatac cacctcagct
     4021 tctcaacgcg tgaagtggtt cggttggtta ggggcaagtt ttgggcttgg tttaatagcg
     4081 gggcctatta ttggtggttt tgcaggagag atttcaccgc atagtccctt ttttatcgct
     4141 gcgttgctaa atattgtcac tttccttgtg gttatgtttt ggttccgtga aaccaaaaat
     4201 acacgtgata atacagatac cgaagtaggg gttgagacgc aatcaaattc ggtgtacatc
     4261 actttattta aaacgatgcc cattttgttg attatttatt tttcagcgca attgataggc
     4321 caaattcccg caacggtgtg ggtgctattt accgaaaatc gttttggatg gaatagcatg
     4381 atggttggct tttcattagc gggtcttggt cttttacact cagtattcca agcctttgtg
     4441 gcaggaagaa tagccactaa atggggcgaa aaaacggcag tactgctcga atttattgca
     4501 gatagtagtg catttgcctt tttagcgttt atatctgaag gttggttaga tttccctgtt
     4561 ttaattttat tggctggtgg tgggatcgct ttacctgcat tacagggagt gatgtctatc
     4621 caaacaaaga gtcatgagca aggtgcttta cagggattat tggtgagcct taccaatgca
     4681 accggtgtta ttggcccatt actgtttact gttatttata atcattcact accaatttgg
     4741 gatggctgga tttggattat tggtttagcg ttttactgta ttattatcct gctatcaatg
     4801 accttcatgt tgacccctca agctcagggg agtaaacagg agacaagtgc ttagttattt
     4861 cgtcaccaaa tgaggcctcc aaaacttcat ttttaatttt tgagctcgct tggactcctg
     4921 ttgatagatc cagtaatgac ctcagaactc catctggatt tgttcagaac gctcggttgc
     4981 cgccgggcgt tttttattgg tgagaatcca agcgtcgacc tagtgcggcc gcaagatccg
     5041 gccacgatgc gtccggcgta gaggatctga agatcagcag ttcaacctgt tgatagtacg
     5101 tactaagctc tcatgtttca cgtactaagc tctcatgttt aacgtactaa gctctcatgt
     5161 ttaacgaact aaaccctcat ggctaacgta ctaagctctc atggctaacg tactaagctc
     5221 tcatgtttca cgtactaagc tctcatgttt gaacaataaa attaatataa atcagcaact
     5281 taaatagcct ctaaggtttt aagttttata agaaaaaaaa gaatatataa ggcttttaaa
     5341 gcttttaagg tttaacggtt gtggacaaca agccagggat gtaacgcact gagaagccct
     5401 tagagcctct caaagcaatt ttgagtgaca caggaacact taacggctga catgggaatt
     5461 aggaagttcc tattccgaag ttcctattct ctagaaagta taggaacttc catatgccat
     5521 ggaatgaccg accgatacaa tgatgatcgt ggtaaagact ggattatcgt agatatcaaa
     5581 acagcataga aaatagccag ttttggggaa gggttttggg gaaagttttg gggaagattt
     5641 tacatcatca taaaacaacg ggcgtgttat acgcccgtta taatatttaa cacatgtggt
     5701 aattacatat ttttgatgat cgcgtcacca aactcgctgc atttcagcag tttagcgcct
     5761 tccatcagac gttcgaagtc ataggttacg gtcttggcgt tgattgcgcc ttccatacct
     5821 ttaacaatca ggtctgcggc ttcagtccag cccatatggc gcagcagtat ataaaaatta
     5881 tgtattcaac tcgctgatat ttaatcataa atattatttt cttgtctttg tattagccaa
     5941 aatcaacttt attcaaccgt ttgatttcat tgacgttcgt ttttattttg gggaaagatt
     6001 ttctctacca tttatatcaa aattacaagt ctacatttta ttaacatctc aatacatgtg
     6061 tggccttgca agccattgcg aggccttatg ctagcttctt cgtctgtttc tactggtatt
     6121 ggcacaaacc tgattccaat ttgagcaagg ctatgtgcca tctcgatact cgttcttaac
     6181 tcaacagaag atgctttgtg catacagccc ctcgtttatt atttatctcc tcagccagcc
     6241 gctgtgcttt cagtggattt cggataacag aaaggccggg aaatacccag cctcgctttg
     6301 taacggagta gagacgaaag tgattgcgcc tacccggata ttatcgtgag gatgcgtcat
     6361 cgccattaat tcactgatca gtgatagctg tcaaacatga gaattgatcc ggctgcctcg
     6421 cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag
     6481 cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg
     6541 gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga tagcggagtg tatgctgcac
     6601 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtcttca agaattaatt
     6661 cccaattcca ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt
     6721 atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa atccgccggg agcggatttg
     6781 aacgttgcga agcaacggcc cggagggtgg cgggcaggac gcccgccata aactgccagg
     6841 aattaattcc aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg cctttcgttt
     6901 tatctgttgt ttgtcggtga acgctctcct gagtaggaca aatccgccgg gagcggattt
     6961 gaacgttgcg aagcaacggc ccggagggtg gcgggcagga cgcccgccat aaactgccag
     7021 gaattaattc caggcatcaa ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt
     7081 ttatctgttg tttgtcggtg aacgctctcc tgagtaggac aaatccgccg ggagcggatt
     7141 tgaacgttgc gaagcaacgg cccggagggt ggcgggcagg acgcccgcca taaactgcca
     7201 ggaattaatt ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt
     7261 tttatctgtt gtttgtcggt gaacgctctc ctgagtagga caaatccgcc gggagcggat
     7321 ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag gacgcccgcc ataaactgcc
     7381 aggaattggg gatcg
//
