LOCUS       62056_21060        8055 bp DNA     circular SYN 06-OCT-2020
DEFINITION  Cloning vector pTcmKateN-NH 2.0, complete sequence.
ACCESSION   MT180307
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8055)
  AUTHORS   Diaz-Olmos Y, Batista M, Ludwig A, Marchini FK.
  TITLE     Characterising ISWI chromatin remodeler in Trypanosoma cruzi
  JOURNAL   Mem. Inst. Oswaldo Cruz 115, e190457 (2020)
  PUBMED    32428081
REFERENCE   2  (bases 1 to 8055)
  AUTHORS   Diaz-Olmos Y, Batista M, Ludwig A, Marchini FK.
  TITLE     Direct Submission
  JOURNAL   Submitted (09-MAR-2020) LACTAS, Fiocruz, Professor Algacyr Munhoz 
            Mader, Curitiba 81350010, Brasil
REFERENCE   3  (bases 1 to 8055)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mem. Inst. 
            Oswaldo Cruz 115, e190457 (2020)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (09-MAR-2020) LACTAS, Fiocruz, Professor Algacyr Munhoz Mader, 
            Curitiba 81350010, Brasil"
FEATURES             Location/Qualifiers
     source          1..8055
                     /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      706..1322
                     /note="Trypanosoma cruzi 18S ribosomal RNA promoter region"
                     /regulatory_class="promoter"
     misc_feature    1335..1839
                     /note="Trypanosoma cruzi 3' intergenic region of the 
                     TcCLB.511469.50 CDS"
     CDS             1849..2643
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase from Tn5"
     misc_feature    2653..2970
                     /note="Trypanosoma cruzi 3' intergenic region of the 
                     TcCLB.511145.40 CDS"
     CDS             2983..3678
                     /label=mKate2
                     /note="monomeric far-red fluorescent protein (Shcherbo et
                     al., 2009)"
     protein_bind    3688..3812
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        3837..3867
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             3921..4577
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     CDS             4922..5224
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
     protein_bind    complement(5268..5392)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    5416..5836
                     /note="Trypanosoma cruzi 3' intergenic region of the 
                     TcCLB.504069.70 CDS"
     promoter        complement(5867..5885)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5906..5922)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(5930..5946)
                     /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(5954..5984)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(5999..6020)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(6308..6896)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(7070..7927)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(7928..8032)
                     /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 tggagctcgc
      661 ggccgcccac ttgtatcgtg ggcgccactt gaatcgtgga tcgatcacca gtttccttgc
      721 ggggcttatt tggtgtgttg gatttttaat tttttgatga ttcggaggaa ttttttgcaa
      781 tttttgggta cccctaaaaa aatttttgga gtttccgggg ttcgatccaa cttgtgcttt
      841 tttaaccgtc catttgtgtc tagtacatca tggagcggta ttctctccgg tggcccaggt
      901 ggcgtcggaa ggaattggtg tccctcagga tgtttggttt ttcgtttttt tttttgcggt
      961 cttttgtttc gcatcgccgt atcgtgtgcg gcgcgacctt ctttttggct cattggtcag
     1021 tgaggtttat aataaagtgg atcatttggg tttttggccc tttgcggcgc acccgcggaa
     1081 tgggggaggg attttcatat aatgtagtgt acattgtgtg tgggtcgcaa aatgtcatgc
     1141 cgtgtgcaat gtttatttgc gtgctgtgcg tgtccttttg tgattccacc acttgtgtgt
     1201 attttacctt atcgtctctc cctgccgttg gctgtgcgtg aattgtgctt cgcattcgtg
     1261 ggaaatcaaa aggggaatca acgtgttctt ttttttcaga aggggagtca aagcgccgtg
     1321 tgatcgatca tatggatttg ggtaggatga gaagggagtt gctgaggaaa aggggaggga
     1381 atcattaata attagaagag ttttttcttc tctctcattt cctgtatcta tatatgtatt
     1441 aggagctgta ttggcaactt ggtggagcat cggcggggtg tttgcggcgc ggacctctgc
     1501 aaacttccaa aggtggtaat gggaatttga aataacttgc caatggacat gactcaacag
     1561 acgaaagcgg tagcaacgat actatgtaat agctgttgtt gttgttattt ttatgtttat
     1621 ttttattatt gatacttcgc ttcggtactc tctctgtttt tttttttctc tcactcattc
     1681 cctctttata tatatatata tatatatata tatatatata tatcgtacgt gtctttttgt
     1741 tgttgttctc aacttcacgt tggctttttt tttttttttt ttgacttgga ttgtatcgga
     1801 cggcccaata acaataataa ttaaaataaa agtggggaaa agcttactag tatgattgaa
     1861 caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac
     1921 tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg
     1981 cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag
     2041 gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt
     2101 gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg
     2161 tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg
     2221 catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga
     2281 gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag
     2341 gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gaatgcccga cggcgatgat
     2401 ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt
     2461 tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg
     2521 gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt
     2581 tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc
     2641 ttctgactcg agcggcatcg gatgcgtttt atggttctgt cttttttttc ccctcctttt
     2701 tttttttctt ttaatttatt ttactttact gcacggtacc ttgtaggaaa tgagatgtgt
     2761 tttttttttt gttttttttg ttgttgttgt tgttgttgag attttttttt tttttttttt
     2821 ttttgatctt acgtgtgtcc tgtgttactg agaatattta tgctgcgcat ttggagatat
     2881 tcatagttgt ttaaatggac tttttctgct gctcaaactt ttgttttctg cgtaattaac
     2941 agggaaggaa actttctaag tcgttcatca gatatctcta gaatggtgag cgagctgatt
     3001 aaggagaaca tgcacatgaa gctgtacatg gagggcaccg tgaacaacca ccacttcaag
     3061 tgcacatccg agggcgaagg caagccctac gagggcaccc agaccatgag aatcaaggcg
     3121 gtcgagggcg gccctctccc cttcgccttc gacatcctgg ctaccagctt catgtacggc
     3181 agcaaaacct tcatcaacca cacccagggc atccccgact tctttaagca gtccttcccc
     3241 gagggcttca catgggagag agtcaccaca tacgaagacg ggggcgtgct gaccgctacc
     3301 caggacacca gcctccagga cggctgcctc atctacaacg tcaagatcag aggggtgaac
     3361 ttcccatcca acggccctgt gatgcagaag aaaacactcg gctgggaggc ctccaccgag
     3421 accctgtacc ccgctgacgg cggcctggaa ggcagagccg acatggccct gaagctcgtg
     3481 ggcgggggcc acctgatctg caacttgaag accacataca gatccaagaa acccgctaag
     3541 aacctcaaga tgcccggcgt ctactatgtg gacagaagac tggaaagaat caaggaggcc
     3601 gacaaagaga cctacgtcga gcagcacgag gtggctgtgg ccagatactg cgacctccct
     3661 agcaaactgg ggcacagagc atgcatcaca agtttgtaca aaaaagctga acgagaaacg
     3721 taaaatgata taaatatcaa tatattaaat tagattttgc ataaaaaaca gactacataa
     3781 tactgtaaaa cacaacatat ccagtcacta tggcggccgc attaggcacc ccaggcttta
     3841 cactttatgc ttccggctcg tataatgtgt ggattttgag ttaggatccg tcgagatttt
     3901 caggagctaa ggaagctaaa atggagaaaa aaatcactgg atataccacc gttgatatat
     3961 cccaatggca tcgtaaagaa cattttgagg catttcagtc agttgctcaa tgtacctata
     4021 accagaccgt tcagctggat attacggcct ttttaaagac cgtaaagaaa aataagcaca
     4081 agttttatcc ggcctttatt cacattcttg cccgcctgat gaatgctcat ccggaattcc
     4141 gtatggcaat gaaagacggt gagctggtga tatgggatag tgttcaccct tgttacaccg
     4201 ttttccatga gcaaactgaa acgttttcat cgctctggag tgaataccac gacgatttcc
     4261 ggcagtttct acacatatat tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt
     4321 tccctaaagg gtttattgag aatatgtttt tcgtctcagc caatccctgg gtgagtttca
     4381 ccagttttga tttaaacgtg gccaatatgg acaacttctt cgcccccgtt ttcaccatgg
     4441 gcaaatatta tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatcatg
     4501 ccgtttgtga tggcttccat gtcggcagaa tgcttaatga attacaacag tactgcgatg
     4561 agtggcaggg cggggcgtaa acgcgtggat ccggcttact aaaagccaga taacagtatg
     4621 cgtatttgcg cgctgatttt tgcggtataa gaatatatac tgatatgtat acccgaagta
     4681 tgtcaaaaag aggtatgcta tgaagcagcg tattacagtg acagttgaca gcgacagcta
     4741 tcagttgctc aaggcatata tgatgtcaat atctccggtc tggtaagcac aaccatgcag
     4801 aatgaagccc gtcgtctgcg tgccgaacgc tggaaagcgg aaaatcagga agggatggct
     4861 gaggtcgccc ggtttattga aatgaacggc tcttttgctg acgagaacag gggctggtga
     4921 aatgcagttt aaggtttaca cctataaaag agagagccgt tatcgtctgt ttgtggatgt
     4981 acagagtgat attattgaca cgcccgggcg acggatggtg atccccctgg ccagtgcacg
     5041 tctgctgtca gataaagtct cccgtgaact ttacccggtg gtgcatatcg gggatgaaag
     5101 ctggcgcatg atgaccaccg atatggccag tgtgccggtc tccgttatcg gggaagaagt
     5161 ggctgatctc agccaccgcg aaaatgacat caaaaacgcc attaacctga tgttctgggg
     5221 aatataaatg tcaggctccc ttatacacag ccagtctgca ggtcgaccat agtgactgga
     5281 tatgttgtgt tttacagtat tatgtagtct gttttttatg caaaatctaa tttaatatat
     5341 tgatatttat atcattttac gtttctcgtt cagctttctt gtacaaagtg gtgatataag
     5401 cggccgcggc gcgcctatgg tagtgaatgg atatatatat atatatatat atatatatat
     5461 aaggtgtaaa tgaaattagt ggaagaggga attagatggg tagcgcgttg cgtggcgctg
     5521 gggagtgtct cgttttatga cttttgtatc tctctgtttg agtgtgtgtg tgtgtgtgtg
     5581 tgtgttttat tctttttttg ttgttgttgt tcttgttaaa tatttattaa tgatttgctt
     5641 ttccttcttg tgtggcttga aattatttcc tctactgttt gtctccacgt caattttgcc
     5701 cataactcat cttgactttc aatccttagc gagtgttttg ttgtttttgg gatagtcgtt
     5761 ttttgtttgt gtgtgtgtgt gtgtgtgtgt gtattttttt cttttttctt cttcttggag
     5821 gggctcctac acgtttaccg gtgggcccgg tacccagctt ttgttccctt tagtgagggt
     5881 taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     5941 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     6001 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     6061 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     6121 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     6181 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     6241 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     6301 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     6361 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     6421 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     6481 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     6541 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     6601 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     6661 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     6721 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     6781 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     6841 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     6901 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     6961 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     7021 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     7081 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     7141 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     7201 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     7261 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     7321 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     7381 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     7441 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     7501 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     7561 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     7621 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     7681 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     7741 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     7801 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     7861 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     7921 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     7981 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     8041 cccgaaaagt gccac
//
