LOCUS       40924_39872        9391 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pSG463, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9391)
  AUTHORS   Goldfless SJ, Wagner JC, Niles JC.
  TITLE     Versatile control of Plasmodium falciparum gene expression with an 
            inducible protein-RNA interaction
  JOURNAL   Nat Commun 5, 5329 (2014)
  PUBMED    25370483
REFERENCE   2  (bases 1 to 9391)
  AUTHORS   Goldfless SJ, Niles JC.
  TITLE     Direct Submission
  JOURNAL   Submitted (10-AUG-2014) Biological Engineering, MIT, 77 
            Massachusetts Avenue, Cambridge, MA 02139, USA
REFERENCE   3  (bases 1 to 9391)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 9391)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat Commun 
            5, 5329 (2014)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (10-AUG-2014) Biological Engineering, MIT, 77 Massachusetts Avenue, 
            Cambridge, MA 02139, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..9391
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     38..54
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        61..79
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_recomb     join(88..138,144..194)
                     /label=tandem Bxb1 attP sites
                     /note="tandem Bxb1 attP sites"
     promoter        complement(230..248)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(266..282)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(290..306)
                     /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(314..344)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(359..380)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(668..1256)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             1845..2651
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
     terminator      2660..2857
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     regulatory      complement(2905..3468)
                     /label=derived from P. falciparum HRPII 3' UTR
                     /note="derived from P. falciparum HRPII 3' UTR"
                     /regulatory_class="terminator"
     CDS             complement(3502..3894)
                     /label=BSD
                     /note="blasticidin S deaminase"
     misc_feature    complement(3901..3963)
                     /note="Thosea asigna virus 2A-like peptide, P. falciparum 
                     optimized codon usage"
     CDS             complement(3901..3954)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             complement(3961..4896)
                     /label=RLuc
                     /note="Renilla luciferase"
     CDS             complement(4897..4950)
                     /label=T2A
                     /note="2A peptide from Thosea asigna virus capsid protein"
     CDS             complement(4960..5580)
                     /label=TetR
                     /note="tetracycline repressor TetR"
     regulatory      complement(5591..6195)
                     /note="promoter and 5'UTR from P. chabaudi dihydrofolate 
                     reductase-thymidylate synthase gene"
                     /regulatory_class="promoter"
     regulatory      6196..6825
                     /note="contains promoter and 5' UTR from P. falciparum 
                     calmodulin gene"
                     /regulatory_class="promoter"
     misc_feature    6828..6874
                     /label=mutant TetR aptamer
                     /note="mutant TetR aptamer"
     CDS             6897..8546
                     /label=luciferase
                     /note="firefly luciferase"
     regulatory      8556..9391
                     /label=derived from P. falciparum hsp86 3' UTR
                     /note="derived from P. falciparum hsp86 3' UTR"
                     /regulatory_class="terminator"
ORIGIN
        1 tgcaggtcga cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgaattg
       61 taatacgact cactataggg cgaattctgg tttgtctggt caaccaccgc ggtctcagtg
      121 gtgtacggta caaacccgaa ttctggtttg tctggtcaac caccgcggtc tcagtggtgt
      181 acggtacaaa cccggaattc gagctcgggc ggccgccaag cttgagtatt ctatagtgtc
      241 acctaaatag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
      301 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
      361 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
      421 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
      481 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
      541 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
      601 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
      661 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
      721 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
      781 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
      841 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
      901 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
      961 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     1021 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     1081 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     1141 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     1201 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     1261 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     1321 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     1381 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     1441 tcagtgaggc agcttcgtac gctgcaggtc gagatttccc cgaaaagtgc cacctgggtc
     1501 cttttcatca cgtgggacat tgtttaaacg agcagggacg tgcgcagctc aggggcatga
     1561 tgtgactgtc gcccgtacat ttagcccata catccccatg tataatcatt tgcatccata
     1621 cattttgatg gccgcacggc gcgaagcaaa aattacggct cctcgctgca gacctgcgag
     1681 cagggaaacg ctcccctcac agacgcgttg aattgtcccc acgccgcgcc cctgtagaga
     1741 aatataaaag gttaggattt gccactgagg ttcttctttc atatacttcc ttttaaaatc
     1801 ttgctaggat acagttctca catcacatcc gaacataaac aaccatgggt aaggaaaaga
     1861 ctcacgtttc gaggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat
     1921 gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgattgtat gggaagcccg
     1981 atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg
     2041 agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta
     2101 tccgtactcc tgatgatgca tggttactca ccactgcgat ccccggcaaa acagcattcc
     2161 aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc
     2221 tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat cgcgtatttc
     2281 gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg
     2341 acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataag cttttgccat
     2401 tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg
     2461 aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg
     2521 atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt
     2581 ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg
     2641 atgagttttt ctaatcagta ctgacaataa aaagattctt gttttcaaga acttgtcatt
     2701 tgtatagttt ttttatattg tagttgttct attttaatca aatgttagcg tgatttatat
     2761 tttttttcgc ctcgacatca tctgcccaga tgcgaagtta agtgcgcaga aagtaatatc
     2821 atgcgtcaat cgtatgtgaa tgctggtcgc tatactgctg tcgattcgat actaacgccg
     2881 ccatccagtg tcgaaaacga gctcggtacc tctagatgta ataaatatgt tcttatatat
     2941 aatgagaaat aaatatttaa catatgttta aaaagaaaaa tttaagattt acatgattag
     3001 gaataaaaat attaaaagct atatattaat gttgtacttt atgttcatat tgtaaaagat
     3061 aaatataatt tttattaaaa tttatattat tattattata agttcttaga aaatacattt
     3121 tgttttaaaa taatactata aaaagaacaa taataaaaat aaaataaata caattaaaac
     3181 ataattaaaa aagaaaatac agacagtaaa aaaaatcgct atcccataaa ttacaaaaca
     3241 tgaattaata aacatatatt gtttattaat aaaattgtaa gttattttta tatatacata
     3301 ataaattaaa gatactgaaa caataatttt aaatgtaaat aaatgaataa tgatatgttt
     3361 tatatgattc attattctat atttataagg aagattacaa aaaaaaaatt catatgtatt
     3421 ttttttgtaa tttctgtgtt tatgttttta tttttataat atttttaatc tattattaaa
     3481 taaatttaat ggaccggttt agccctccca cacataacca gagggcagca attcacgaat
     3541 cccaactgcc gtcggctgtc catcactgtc cttcactatg gctttgatcc caggatgcag
     3601 atcgagaagc acctgtcggc accgtccgca ggggctcaag atgcccctgt tctcatttcc
     3661 gatcgcgacg atacaagtca ggttgccagc tgccgcagca gcagcagtgc ccagcaccac
     3721 gagttctgca caaggtcccc cagtaaaatg atatacattg acaccagtga agatgcggcc
     3781 gtcgctagag agagctgcgc tggcgacgct gtagtcttca gagatgggga tgctgttgat
     3841 tgtagccgtt gctctttcaa tgagggtgga ttcttcttga gacaaaggct tggcgccggc
     3901 tggtcctgga ttttcttcta catctccaca tgttaataaa cttcctcttc cttctccact
     3961 tccttgttca tttttgagaa ctcgctcaac gaacgatttg atatattttc ccatttcatc
     4021 aggtgcatct tcttgcgaaa aatgaagacc ttttactttg acaaattcag tattaggaaa
     4081 cttcttggca ccttcaacaa tagcattgga aaagaatcct gggtccgatt caataaacat
     4141 ttttggtaaa tcatcacttg cacgtagata agcattataa ttcctaacaa tttgtacaac
     4201 gtcaggttta ccacctttta ctaacgggat ttcacgaggc catgataatg ttggacgacg
     4261 aacttcacct ttctctttga atggttcaag atatgctgca aattcttctg gttctaactt
     4321 tctcatgatt tttgatggca acatggtttc cacgaagaag ttattctcca aaaccatttt
     4381 ttctccttct tcagatttga tcaacgcaat atcttcttca atatcaggcc attcatccca
     4441 tgattcaatc acatctacta cactttcagc gtgaactatt gctttgatct tatcttgatg
     4501 ctcatagcta taatgaaatg ccaaacaagc accccaatca tggccgacaa aaatgatctt
     4561 ctttggtaaa ttaagaagtt caaaccatgc agtaagatat ttgtaatgat caagtaacct
     4621 ataagaacca ttaccagatt tgcctgattt gcccatacca ataaggtctg gtataataca
     4681 ccgcgctact ggctcaatat gtggcacaac atgtcgccat aaataagaag aggccgcgtt
     4741 accatgtaaa aaaataacag cattttctgc atgtttttct gaatcataat aattaataaa
     4801 tgaatcaaga acattcattt gtttacatct ggcccaccac tgcggaccag ttatcatccg
     4861 tttcctttgt tctggatcat aaactttcga agtcattgga cctggatttt cttcaacatc
     4921 accacatgtt aataatgatc ctctaccttc accactacca gacccacttt cacatttaag
     4981 ttgtttttct aatccgcata tgatcaattc aaggccgaat aagaaggctg gctctgcacc
     5041 ttggtgatca aataattcga tagcttgtcg taataatggc ggcatactat cagtagtagg
     5101 tgtttccctt tcttctttag cgacttgatg ctcttgatct tccaatacgc aacctaaggt
     5161 aaaatgcccc acagcgctga gtgcatataa cgcgttctct agtgaaaaac cttgttggca
     5221 taaaaaggct aattgatttt cgagagtttc atactgtttt tctgtaggcc gtgtacctaa
     5281 atgtactttt gctccatcgc gatgacttag taaagcacat ctaaaacttt tagcgttatt
     5341 acgtaaaaaa tcttgccagc tttccccttc taaagggcag aagtgagtat ggtgcctatc
     5401 taacatctca atggctaagg cgtcgagcaa agcccgctta ttttttacat gccaatacaa
     5461 tgtaggctgc tctacaccaa gcttctgggc gagtttacgg gttgttaaac cttcgattcc
     5521 gacctcatta agcagctcta atgcgctgtt aatcacttta cttttatcta atctagacat
     5581 ttttcctagg tttgtaagtt ttaggtgtgt gtatatattt aagcactaca caattgttac
     5641 tagcaaatgt agaaagaaat agccagtatt taaactaaat atgtgaatat taaaatatag
     5701 tgacaaaaaa ggaaagtgta tgtaaaataa aatgaaaatt gaaaagaatt tatattataa
     5761 tataaaaaat tgtagtgaac atatataatc ctgtgttata tatatttaat attattattg
     5821 acttttttaa aaaaaatatt tattataaaa attttttaat ttatgcactt ttttattgta
     5881 cttatatata ttccattatt ttacttacac ctaaatttag tagaaaattt tattcaaccc
     5941 aaataaaaat tatggcgcta tataggatga gatcatatat ttttttctct ttttcaaatc
     6001 ctttatattt tatcatacaa actgttttaa acacgagaaa aaggggatgc catgcacatg
     6061 cttagtacac atgcaataga tatatatagt atatatatta catatatata tgtgtacttg
     6121 ttaaaatgct atatcaaata gttttgcata aacgagagta cctcttacaa gcgaattagc
     6181 taacctagtg tttataagct tttatatgtg attaatttta tatattatca atttatatat
     6241 ttttaaatgc ttacttaatt atcttttttt tttttttttt ttttttttcc cctcttttta
     6301 tattaattta tttttgaaaa aattgatata tatatatata tataatatat atatatacat
     6361 gtagtagtat taaacaatgt ataatatata taaataatat atttatatat ttcatttcaa
     6421 ttttaatttt ttttggtttt tttttttttt ctttttgtca tatttaaaaa aaattatatt
     6481 catataagtt atgcattttt tataaacatt attcaatata tgtataatat aatatatata
     6541 tatatattaa tgtattattc caatgtgcat gataaaagaa aaaaataata tttataaaaa
     6601 aaaagaaaaa taaaacaaaa aaagaaaaaa aaaaaaaaaa aaaaaaaaat acaaaaataa
     6661 ataatataat ttataattat atattcttgt cacaataaaa atatatatat atatatatat
     6721 atttataata tgtatatttt aaactagaaa aggaataact aatattttat ttattatcat
     6781 tcaagattta tattttataa taataaatac ctaatagaaa tatatcaatc caggcagtgt
     6841 aaggtcgata cggacggaat gtgatggcct ggataaaaca caaaactcga gaacatatgg
     6901 aagacgccaa aaacataaag aaaggcccgg cgccattcta tccactagag gatggaaccg
     6961 ctggagagca actgcataag gctatgaaga gatacgccct ggttcctgga acaattgctt
     7021 ttacagatgc acatatcgag gtgaacatca cgtacgcgga atacttcgaa atgtccgttc
     7081 ggttggcaga agctatgaaa cgatatgggc tgaatacaaa tcacagaatc gtcgtatgca
     7141 gtgaaaactc tcttcaattc tttatgccgg tgttgggcgc gttatttatc ggagttgcag
     7201 ttgcgcccgc gaacgacatt tataatgaac gtgaattgct caacagtatg aacatttcgc
     7261 agcctaccgt agtgtttgtt tccaaaaagg ggttgcaaaa aattttgaac gtgcaaaaaa
     7321 aattaccaat aatccagaaa attattatca tggattctaa aacggattac cagggatttc
     7381 agtcgatgta cacgttcgtc acatctcatc tacctcccgg ttttaatgaa tacgattttg
     7441 taccagagtc ctttgatcgt gacaaaacaa ttgcactgat aatgaattcc tctggatcta
     7501 ctgggttacc taagggtgtg gcccttccgc atagaactgc ctgcgtcaga ttctcgcatg
     7561 ccagagatcc tatttttggc aatcaaatca ttccggatac tgcgatttta agtgttgttc
     7621 cattccatca cggttttgga atgtttacta cactcggata tttgatatgt ggatttcgag
     7681 tcgtcttaat gtatagattt gaagaagagc tgtttttacg atcccttcag gattacaaaa
     7741 ttcaaagtgc gttgctagta ccaaccctat tttcattctt cgccaaaagc actctgattg
     7801 acaaatacga tttatctaat ttacacgaaa ttgcttctgg gggcgcacct ctttcgaaag
     7861 aagtcgggga agcggttgca aaacgcttcc atcttccagg gatacgacaa ggatatgggc
     7921 tcactgagac tacatcagct attctgatta cacccgaggg ggatgataaa ccgggcgcgg
     7981 tcggtaaagt tgttccattt tttgaagcga aggttgtgga tctggatacc gggaaaacgc
     8041 tgggcgttaa tcagagaggc gaattatgtg tcagaggacc tatgattatg tccggttatg
     8101 taaacaatcc ggaagcgacc aacgccttga ttgacaagga tggatggcta cattctggag
     8161 acatagctta ctgggacgaa gacgaacact tcttcatagt tgaccgcttg aagtctttaa
     8221 ttaaatacaa aggatatcag gtggcccccg ctgaattgga atcgatattg ttacaacacc
     8281 ccaacatctt cgacgcgggc gtggcaggtc ttcccgacga tgacgccggt gaacttcccg
     8341 ccgccgttgt tgttttggag cacggaaaga cgatgacgga aaaagagatc gtggattacg
     8401 tcgccagtca agtaacaacc gcgaaaaagt tgcgcggagg agttgtgttt gtggacgaag
     8461 taccgaaagg tcttaccgga aaactcgacg caagaaaaat cagagagatc ctcataaagg
     8521 ccaagaaggg cggaaagtcc gagttgtaac tcgagttata taatatattt atgtactcac
     8581 aatggggtct acaaaaaaaa aaaaataaag atatatatat atatatatta tctttcattt
     8641 tatcatcacc ccaatataca tatatatatt tttatatttt tatatattta tatttttctt
     8701 ttatttatcc ataaaagaaa attaaatata aatttatacc ttttatttga attaccattt
     8761 gtataaaata taaaaaaaaa aaaaaataat aaaaagaaga aaaaacgaaa aaagtaatat
     8821 ttaaataata aagatcaata tacataataa aaaaaaaaaa aaaaaaaaaa attaaaattt
     8881 aaaaaattaa acattgaaat aatttcatat acacacatat acatacatac atatatatat
     8941 atatatatat atatttatac atttatgcat ataaaatata ttaacaactt atgtgtattc
     9001 atatatattt ttttacatat gtattgaaag actttactga gacatgaaat atctaataac
     9061 caaacgttta taaattctaa ttatatgtaa agcataatat agcttttgtc taatttaatt
     9121 ccttaaaacg tatgtattaa aaatatagta agatggggaa attataattt tttttttaaa
     9181 ggtttttctt tgatacagta aaaaaaaaaa ataaaaaaaa ataataataa taaaaaaata
     9241 aaaaaataaa aaaaaataaa ataattataa agatatatat aattcctccc aaatacaaat
     9301 gattaagttt aaaaattaat aaataaatca aaaaaaaatt taataataat tgtattattt
     9361 taagtgtagt taattcatca aatagcatgc c
//
