LOCUS       40924_4754        5004 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pARKANI, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5004)
  AUTHORS   Eguia FAP., Ramos HR, Kraschowetz S, Omote DQ, Ramos CRR., Ho PL, 
            Carvalho E, Goncalves VM.
  TITLE     A new vector for heterologous gene expression in Escherichia coli 
            with increased stability in the absence of antibiotic
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5004)
  AUTHORS   Eguia FAP., Ramos HR, Kraschowetz S, Omote DQ, Ramos CRR., Ho PL, 
            Carvalho E, Goncalves VM.
  TITLE     Direct Submission
  JOURNAL   Submitted (06-OCT-2017) Centro de Biotecnologia, Instituto Butantan,
            Av Vital Brasil, 1500, Sao Paulo, SP 05503-900, Brazil
REFERENCE   3  (bases 1 to 5004)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5004)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (06-OCT-2017) Centro de Biotecnologia, Instituto Butantan, Av Vital 
            Brasil, 1500, Sao Paulo, SP 05503-900, Brazil"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Assembly Method       :: CLC Genomics Workbench v. 9.0
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..5004
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             complement(11..33)
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(48..72)
                     /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(73..91)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        400..477
                     /label=lacI promoter
     CDS             601..969
                     /codon_start=1
                     /gene="lacI"
                     /product="Lac repressor"
                     /label=lacI
                     /protein_id="AVN68413.1"
                     /translation="MAELNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSR
                     ADQLGASVVVSMVERSGVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPA
                     LFLDVSDQTPINSIIFLP"
     gene            601..969
                     /gene="lacI"
                     /label=lacI
     protein_bind    1574..1595
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    complement(1946..2314)
                     /label=plasmid partitioning sequence - par
                     /note="plasmid partitioning sequence - par"
     primer_bind     complement(2356..2372)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2380..2396)
                     /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(2404..2434)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(2449..2470)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(2759..3349)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3637..4428)
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     terminator      complement(4823..4870)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(4987..5004)
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
ORIGIN
        1 catatgtata tctccttctt aaagttaaac aaaattattt ctagagggga attgttatcc
       61 gctcacaatt cccctatagt gagtcgtatt aatttcgcgg gatcgagatc tcgatcctct
      121 acgccggacg catcgtggcc ggcatcaccg gcgccacagg tgcggttgct ggcgcctata
      181 tcgccgacat caccgatggg gaagatcggg ctcgccactt cgggctcatg agcgcttgtt
      241 tcggcgtggg tatggtggca ggccccgtgg ccgggggact gttgggcgcc atctccttgc
      301 atgcaccatt ccttgcggcg gcggtgctca acggcctcaa cctactactg ggctgcttcc
      361 taatgcagga gtcgcataag ggagagcgtc gagatcccgg acaccatcga atggcgcaaa
      421 acctttcgcg gtatggcatg atagcgcccg gaagagagtc aattcagggt ggtgaatgtg
      481 aaaccagtaa cgttatacga tgtcgcagag tatgccggtg tctcttatca gaccgtttcc
      541 cgcgtggtga accaggccag ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg
      601 atggcggagc tgaattacat tcccaaccgc gtggcacaac aactggcggg caaacagtcg
      661 ttgctgattg gcgttgccac ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg
      721 gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga
      781 agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg
      841 ctgatcatta actatccgct ggatgaccag gatgccattg ctgtggaagc tgcctgcact
      901 aatgttccgg cgttatttct tgatgtctct gaccagacac ccatcaacag tattattttt
      961 ctcccatgaa gacggtacgc gactgggcgt ggagcatctg gtcgcattgg gtcaccagca
     1021 aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc tggctggctg
     1081 gcataaatat ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag
     1141 tgccatgtcc ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc
     1201 gatgctggtt gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg
     1261 gctgcgcgtt ggtgcggaca tctcggtagt gggatacgac gataccgaag acagctcatg
     1321 ttatatcccg ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt
     1381 ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt
     1441 ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct ctccccgcgc
     1501 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     1561 agcgcaacgc aattaatgta agttagctca ctcattaggc accgggatct cgaccgatgc
     1621 ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg actatcgtcg
     1681 ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg gcagcgctct
     1741 gggtcatttt cggcgaggac cgctttcgct ggagcgcgac gatgatcggc ctgtcgcttg
     1801 cggtattcgg aatcttgcac gccctcgctc aagccttcgt cactggtccc gccaccaaac
     1861 gtttcggcga gaagcaggcc attatcgccg gcatggcggc cccacgggtg cgcatgatcg
     1921 tgctcctgtc gttgaggacc gatcacgggc aaatcgctga atattccttt tgtctccgac
     1981 catcaggcac ctgagtcgct gtctttttcg tgacattcag ttcgctgcgc tcacggctct
     2041 ggcagtgaat gggggtaaat ggcactacag gcgcctttta tggattcatg caaggaaact
     2101 acccataata caagaaaagc ccgtcacggg cttctcaggg cgttttatgg cgggtctgct
     2161 atgtggtgct atctgacttt ttgctgttca gcagttcctg ccctctgatt ttccagtctg
     2221 accacttcgg attatcccgt gacaggtcat tcagactggc taatgcaccc agtaaggcag
     2281 cggtatcatc aacaggctta cccgtcttac tgtcgaattc aatcgaattc ccgcggccag
     2341 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     2401 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     2461 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     2521 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     2581 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     2641 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     2701 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     2761 tccataggct ccgccccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     2821 cgaaacccga caggactata aagataccag gcgtttcccc cctggaagct ccctcgtgcg
     2881 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     2941 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     3001 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     3061 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     3121 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     3181 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     3241 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     3301 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     3361 gatcttttct acgggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     3421 tcatgagatt atcaaaaagg atcttcacct agatcctttt ggggtgggcg aagaactcca
     3481 gcatgagatc cccgcgctgg aggatcatcc agccggcgtc ccggaaaacg attccgaagc
     3541 ccaacctttc atagaaggcg gcggtggaat cgaaatctcg tgatggcagg ttgggcgtcg
     3601 cttggtcggt catttcgaac cccagagtcc cgctcagaag aactcgtcaa gaaggcgata
     3661 gaaggcgatg cgctgcgaat cgggagcggc gataccgtaa agcacgagga agcggtcagc
     3721 ccattcgccg ccaagctctt cagcaatatc acgggtagcc aacgctatgt cctgatagcg
     3781 gtccgccaca cccagccggc cacagtcgat gaatccagaa aagcggccat tttccaccat
     3841 gatattcggc aagcaggcat cgccatgggt cacgacgaga tcctcgccgt cgggcatccg
     3901 cgccttgagc ctggcgaaca gttcggctgg cgcgagcccc tgatgctctt cgtccagatc
     3961 atcctgatcg acaagaccgg cttccatccg agtacgtgct cgctcgatgc gatgtttcgc
     4021 ttggtggtcg aatgggcagg tagccggatc aagcgtatgc agccgccgca ttgcatcagc
     4081 catgatggat actttctcgg caggagcaag gtgagatgac aggagatcct gccccggcac
     4141 ttcgcccaat agcagccagt cccttcccgc ttcagtgaca acgtcgagca cagctgcgca
     4201 aggaacgccc gtcgtggcca gccacgatag ccgcgctgcc tcgtcttgga gttcattcag
     4261 ggcaccggac aggtcggtct tgacaaaaag aaccgggcgc ccctgcgctg acagccggaa
     4321 cacggcggca tcagagcagc cgattgtctg ttgtgcccag tcatagccga atagcctctc
     4381 cacccaagcg gccggagaac ctgcgtgcaa tccatcttgt tcaatcatgc gaaacgatcc
     4441 tcatcctgtc tcttgatcag atcttgatcc cctgcgccat cagatccttg gcggcaagaa
     4501 agccatccag tttactttgc agggcttccc aaccttacca gagggcgccc cagctggcaa
     4561 ttccggttcg cttgctgtcc ataaaaccgc ccagtctagc tatcgccatg taagcccact
     4621 gcaagctacc tgctttctct ttgcgcttgc gttttccctt gtccagatag cccagtagct
     4681 gacattcatc cggggtcagc accgtttctg cggactggct ttctacgtgt tccgcttcct
     4741 ttagcagccc ttgcgccctg agtgcttgcg gcagcgtgaa gctagtgatt ctcgacgatc
     4801 cggatatagt tcctcctttc agcaaaaaac ccctcaagac ccgtttagag gccccaaggg
     4861 gttatgctag ttattgctca gcggtggcag cagccaactc agcttccttt cgggctttgt
     4921 tagcagccgg atcaagcttc gaattccatg gtaccagctg cagatctcga ggtcggatcc
     4981 ctcgaggtga tggtgatggt gatg
//
