LOCUS       40924_42759        7584 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pTcGFPN, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7584)
  AUTHORS   Batista M, Marchini FK, Celedon PA, Fragoso SP, Probst CM, Preti H, 
            Ozaki LS, Buck GA, Goldenberg S, Krieger MA.
  TITLE     A high-throughput cloning system for reverse genetics in Trypanosoma
            cruzi
  JOURNAL   BMC Microbiol. 10 (1), 259 (2010)
  PUBMED    20942965
REFERENCE   2  (bases 1 to 7584)
  AUTHORS   Batista M, Marchini FK, Celedon PAF., Fragoso SP, Probst CM, Preti 
            H, Ozaki LS, Buck GA, Goldenberg S, Krieger MA.
  TITLE     Direct Submission
  JOURNAL   Submitted (28-APR-2010) Instituto Carlos Chagas, Professor Algacyr 
            Munhoz Mader, 3775, Curitiba, Parana 81350010, Brasil
REFERENCE   3  (bases 1 to 7584)
  AUTHORS   Batista M, Marchini FK, Celedon PAF., Fragoso SP, Probst CM, Preti 
            H, Ozaki LS, Buck GA, Goldenberg S, Krieger MA.
  TITLE     Direct Submission
  JOURNAL   Submitted (18-NOV-2010) Instituto Carlos Chagas, Professor Algacyr 
            Munhoz Mader, 3775, Curitiba, Parana 81350010, Brasil
REFERENCE   4  (bases 1 to 7584)
  TITLE     Direct Submission
REFERENCE   5  (bases 1 to 7584)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "BMC 
            Microbiol."; date: "2010"; volume: "10"; issue: "1"; pages: "259"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (28-APR-2010) Instituto Carlos Chagas, Professor Algacyr Munhoz 
            Mader, 3775, Curitiba, Parana 81350010, Brasil"
COMMENT     SGRef: number: 3; type: "Journal Article"; journalName: "Submitted 
            (18-NOV-2010) Instituto Carlos Chagas, Professor Algacyr Munhoz 
            Mader, 3775, Curitiba, Parana 81350010, Brasil"
COMMENT     SGRef: number: 4; type: "Journal Article"
COMMENT     On Nov 18, 2010 this sequence version replaced HM162843.1.
FEATURES             Location/Qualifiers
     source          1..7584
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(3..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      659..1275
                     /note="derived from Trypanosoma cruzi 18S ribosomal RNA
                     gene promoter region"
                     /regulatory_class="promoter"
     CDS             1569..2282
                     /label=EGFP
                     /note="enhanced GFP"
     protein_bind    2292..2411
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        2441..2471
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             2525..3181
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     CDS             3526..3828
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
     protein_bind    complement(3872..3996)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             4296..5084
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase from Tn5"
     promoter        complement(5396..5414)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5435..5451)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(5459..5475)
                     /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)."
     promoter        complement(5483..5513)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(5528..5549)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(5837..6425)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6599..7456)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(7457..7561)
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggagctcca
      661 ccagtttcct tgcggggctt atttggtgtg ttggattttt aattttttga tgattcggag
      721 gaattttttg caatttttgg gtacccctaa aaaaattttt ggagtttccg gggttcgatc
      781 caacttgtgc ttttttaacc gtccatttgt gtctagtaca tcatggagcg gtattctctc
      841 cggtggccca ggtggcgtcg gaaggaattg gtgtccctca ggatgtttgg tttttcgttt
      901 ttttttttgc ggtcttttgt ttcgcatcgc cgtatcgtgt gcggcgcgac cttctttttg
      961 gctcattggt cagtgaggtt tataataaag tggatcattt gggtttttgg ccctttgcgg
     1021 cgcacccgcg gaatggggga gggattttca tataatgtag tgtacattgt gtgtgggtcg
     1081 caaaatgtca tgccgtgtgc aatgtttatt tgcgtgctgt gcgtgtcctt ttgtgattcc
     1141 accacttgtg tgtattttac cttatcgtct ctccctgccg ttggctgtgc gtgaattgtg
     1201 cttcgcattc gtgggaaatc aaaaggggaa tcaacgtgtt cttttttttc agaaggggag
     1261 tcaaagcgcc gtgtggatat ctctgcgatg ctgtggaccg acgcattgaa atacacaccg
     1321 tctttggcgt tccttttttt ttttttgatt tgtttttttt attgagatgt tttgttttgt
     1381 ttttttcagt ttttattata tcagcagttt gtccgcctgc attcatgcag tgtttggtaa
     1441 ttctttctat tttttggaat tatggcgata atattcttgt ctttaaactt cttataacca
     1501 attgtgcttt agagtttcct gcttagttgc tattaacaca ctgttaggaa cgtgaaacca
     1561 tggcatgcat gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     1621 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     1681 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     1741 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     1801 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     1861 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     1921 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     1981 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     2041 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     2101 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     2161 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     2221 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     2281 agtctagaat cacaagtttg tacaaaaaag ctgaacgaga aacgtaaaat gatataaata
     2341 tcaatatatt aaattagatt ttgcataaaa aacagactac ataatactgt aaaacacaac
     2401 atatccagtc atattggcgg ccgcattagg caccccaggc tttacacttt atgcttccgg
     2461 ctcgtataat gtgtggattt tgagttagga tccgtcgaga ttttcaggag ctaaggaagc
     2521 taaaatggag aaaaaaatca ctggatatac caccgttgat atatcccaat ggcatcgtaa
     2581 agaacatttt gaggcatttc agtcagttgc tcaatgtacc tataaccaga ccgttcagct
     2641 ggatattacg gcctttttaa agaccgtaaa gaaaaataag cacaagtttt atccggcctt
     2701 tattcacatt cttgcccgcc tgatgaatgc tcatccggaa ttccgtatgg caatgaaaga
     2761 cggtgagctg gtgatatggg atagtgttca cccttgttac accgttttcc atgagcaaac
     2821 tgaaacgttt tcatcgctct ggagtgaata ccacgacgat ttccggcagt ttctacacat
     2881 atattcgcaa gatgtggcgt gttacggtga aaacctggcc tatttcccta aagggtttat
     2941 tgagaatatg tttttcgtct cagccaatcc ctgggtgagt ttcaccagtt ttgatttaaa
     3001 cgtggccaat atggacaact tcttcgcccc cgttttcacc atgggcaaat attatacgca
     3061 aggcgacaag gtgctgatgc cgctggcgat tcaggttcat catgccgttt gtgatggctt
     3121 ccatgtcggc agaatgctta atgaattaca acagtactgc gatgagtggc agggcggggc
     3181 gtaaacgcgt ggatccggct tactaaaagc cagataacag tatgcgtatt tgcgcgctga
     3241 tttttgcggt ataagaatat atactgatat gtatacccga agtatgtcaa aaagaggtat
     3301 gctatgaagc agcgtattac agtgacagtt gacagcgaca gctatcagtt gctcaaggca
     3361 tatatgatgt caatatctcc ggtctggtaa gcacaaccat gcagaatgaa gcccgtcgtc
     3421 tgcgtgccga acgctggaaa gcggaaaatc aggaagggat ggctgaggtc gcccggttta
     3481 ttgaaatgaa cggctctttt gctgacgaga acaggggctg gtgaaatgca gtttaaggtt
     3541 tacacctata aaagagagag ccgttatcgt ctgtttgtgg atgtacagag tgatattatt
     3601 gacacgcccg ggcgacggat ggtgatcccc ctggccagtg cacgtctgct gtcagataaa
     3661 gtctcccgtg aactttaccc ggtggtgcat atcggggatg aaagctggcg catgatgacc
     3721 accgatatgg ccagtgtgcc ggtctccgtt atcggggaag aagtggctga tctcagccac
     3781 cgcgaaaatg acatcaaaaa cgccattaac ctgatgttct ggggaatata aatgtcaggc
     3841 tcccttatac acagccagtc tgcaggtcga ccatagtgac tggatatgtt gtgttttaca
     3901 gcattatgta gtctgttttt tatgcaaaat ctaatttaat atattgatat ttatatcatt
     3961 ttacgtttct cgttcagctt tcttgtacaa agtggtgata ctagttagtc tgcgatgctg
     4021 tggaccgacg cattgaaata cacaccgtct ttggcgttcc tttttttttt tttgatttgt
     4081 tttttttatt gagatgtttt gttttgtttt tttcagtttt tattatatca gcagtttgtc
     4141 cgcctgcatt catgcagtgt ttggtaattc tttctatttt ttggaattat ggcgataata
     4201 ttcttgtctt taaacttctt ataaccaatt gtgctttaga gtttcctgct tagttgctat
     4261 taacacactg ttaggaacgt gaaaccctcg agatgattga acaagatgga ttgcacgcag
     4321 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
     4381 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
     4441 agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc
     4501 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
     4561 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
     4621 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     4681 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     4741 ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac
     4801 tgttcgccag gctcaaggcg cgaatgcccg acggcgatga tctcgtcgtg acccatggcg
     4861 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     4921 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     4981 aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     5041 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgactc gagtctgcga
     5101 tgctgtggac cgacgcattg aaatacacac cgtctttggc gttccttttt ttttttttga
     5161 tttgtttttt ttattgagat gttttgtttt gtttttttca gtttttatta tatcagcagt
     5221 ttgtccgcct gcattcatgc agtgtttggt aattctttct attttttgga attatggcga
     5281 taatattctt gtctttaaac ttcttataac caattgtgct ttagagtttc ctgcttagtt
     5341 gctattaaca cactgttagg aacgtgaaac cgggcccggt acccagcttt tgttcccttt
     5401 agtgagggtt aattgcgcgc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     5461 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     5521 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     5581 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     5641 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     5701 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     5761 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     5821 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     5881 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     5941 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     6001 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     6061 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     6121 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     6181 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     6241 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     6301 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     6361 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6421 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     6481 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     6541 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     6601 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6661 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     6721 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     6781 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     6841 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     6901 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     6961 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     7021 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     7081 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     7141 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     7201 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     7261 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     7321 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     7381 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     7441 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     7501 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     7561 gcacatttcc ccgaaaagtg ccac
//
