LOCUS       sequence-148311-        7338 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Directed Evolution of Protein-Protein Interactions.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7338)
  AUTHORS   Badran AH, Guzov VM, Huai Q, Kemp MM, Vishwanath P, Kain W, Nance 
            AM, Evdokimov A, Moshiri F, Turner KH, Wang P, Malvar T, Liu DR
  TITLE     Continuous evolution of Bacillus thuringiensis toxins overcomes 
            insect resistance.
  JOURNAL   Nature. 2016 May 5;533(7601):58-63. doi: 10.1038/nature17938. Epub 
            2016 Apr 27.
  PUBMED    27120167
REFERENCE   2  (bases 1 to 7338)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/nature17938"; journalName: "Nature"; date: "2016-05-5- 5"; 
            volume: "533"; issue: "7601"; pages: "58-63"
COMMENT     Sequence Label: pAB035a
FEATURES             Location/Qualifiers
     source          1..7338
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(1..951)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      999..1221
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     promoter        join(1818..1823,1824..1841,1842..1848)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     CDS             1888..3162
                     /codon_start=1
                     /gene="III"
                     /product="pIII"
                     /label=M13 gene III
                     /note="minor coat protein"
                     /translation="MKKLLFAIPLVVPFYSHSAETVESCLAKPHTENSFTNVWKDDKTL
                     DRYANYEGCLWNATGVVVCTGDETQCYGTWVPIGLAIPENEGGGSEGGGSEGGGSEGGG
                     TKPPEYGDTPIPGYTYINPLDGTYPPGTEQNPANPNPSLEESQPLNTFMFQNNRFRNRQ
                     GALTVYTGTVTQGTDPVKTYYQYTPVSSKAMYDAYWNGKFRDCAFHSGFNEDPFVCEYQ
                     GQSSDLPQPPVNAGGGSGGGSGGGSEGGGSEGGGSEGGGSEGGGSGGGSGSGDFDYEKM
                     ANANKGAMTENADENALQSDAKGKLDSVATDYGAAIDGFIGDVSGLANGNGATGDFAGS
                     NSQMAQVGDGDNSPLMNNFRQYLPSLPQSVECRPFVFGAGKPYEFSIDCDKINLFRGVF
                     AFLLYVATFMYVFSTFANILRNKES"
     CDS             3162..4241
                     /codon_start=1
                     /gene="luxA"
                     /product="LuxA luciferase subunit"
                     /label=LuxA
                     /note="from the Photorhabdus luminescens bioluminescence 
                     operon"
                     /translation="MKFGNFLLTYQPPQFSQTEVMKRLVKLGRISEECGFDTVWLLEHH
                     FTEFGLLGNPYVAAAYLLGATKKLNVGTAAIVLPTAHPVRQLEDVNLLDQMSKGRFRFG
                     ICRGLYNKDFRVFGTDMNNSRALAECWYGLIKNGMTEGYMEADNEHIKFHKVKVNPAAY
                     SRGGAPVYVVAESASTTEWAAQFGLPMILSWIINTNEKKAQLELYNEVAQEYGHDIHNI
                     DHCLSYITSVDHDSIKAKEICRKFLGHWYDSYVNATTIFDDSDQTRGYDFNKGQWRDFV
                     LKGHKDTNRRIDYSYEINPVGTPQECIDIIQKDIDATGISNICCGFEANGTVDEIIASM
                     KLFQSDVMPFLKEKQRSLLY"
     CDS             4305..5285
                     /codon_start=1
                     /gene="luxB"
                     /product="LuxB luciferase subunit"
                     /label=LuxB
                     /note="from the Photorhabdus luminescens bioluminescence 
                     operon"
                     /translation="KFGLFFLNFINSTTVQEQSIVRMQEITEYVDKLNFEQILVYENHF
                     SDNGVVGAPLTVSGFLLGLTEKIKIGSLNHIITTHHPVRIAEEACLLDQLSEGRFILGF
                     SDCEKKDEMHFFNRPVEYQQQLFEECYEIINDALTTGYCNPDNDFYSFPKISVNPHAYT
                     PGGPRKYVTATSHHIVEWAAKKGIPLIFKWDDSNDVRYEYAERYKAVADKYDVDLSEID
                     HQLMILVNYNEDSNKAKQETRAFISDYVLEMHPNENFENKLEEIIAENAVGNYTECITA
                     AKLAIEKCGAKSVLLSFEPMNDLMSQKNVINIVDDNIKKYHTEYT"
     CDS             complement(5693..5722)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(5732..6004)
                     /codon_start=1
                     /gene="E. coli rpoZ"
                     /product="DNA-directed RNA polymerase subunit omega"
                     /label=RNAP omega
                     /note="promotes RNA polymerase assembly"
                     /translation="MARVTVQDAVEKIGNRFDLVLVAARRARQMQVGGKDPLVPEENDK
                     TTVIALREIEEGLINNQILDVRERQEQQEQEAAELQAVTAIAEGRR"
     promoter        6068..6122
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    6069..6087
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6099..6117
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    6123..6141
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             complement(join(6267..7058,7059..7127))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6890..6909
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7128..7232)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7300..7318
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg gttcgttgtt tttgcgtgag
       61 ccatgagaac gaaccattga gatcatgctt actttgcatg tcactcaaaa attttgcctc
      121 aaaactggtg agctgaattt ttgcagttaa agcatcgtgt agtgtttttc ttagtccgtt
      181 acgtaggtag gaatctgatg taatggttgt tggtattttg tcaccattca tttttatctg
      241 gttgttctca agttcggtta cgagatccat ttgtctatct agttcaactt ggaaaatcaa
      301 cgtatcagtc gggcggcctc gcttatcaac caccaatttc atattgctgt aagtgtttaa
      361 atctttactt attggtttca aaacccattg gttaagcctt ttaaactcat ggtagttatt
      421 ttcaagcatt aacatgaact taaattcatc aaggctaatc tctatatttg ccttgtgagt
      481 tttcttttgt gttagttctt ttaataacca ctcataaatc ctcatagagt atttgttttc
      541 aaaagactta acatgttcca gattatattt tatgaatttt tttaactgga aaagataagg
      601 caatatctct tcactaaaaa ctaattctaa tttttcgctt gagaacttgg catagtttgt
      661 ccactggaaa atctcaaagc ctttaaccaa aggattcctg atttccacag ttctcgtcat
      721 cagctctctg gttgctttag ctaatacacc ataagcattt tccctactga tgttcatcat
      781 ctgagcgtat tggttataag tgaacgatac cgtccgttct ttccttgtag ggttttcaat
      841 cgtggggttg agtagtgcca cacagcataa aattagcttg gtttcatgct ccgttaagtc
      901 atagcgacta atcgctagtt catttgcttt gaaaacaact aattcagaca tacatctcaa
      961 ttggtctagg tgattttaat cactatacca attgagatgg gctagtcaat gataattact
     1021 agtccttttc ctttgagttg tgggtatctg taaattctgc tagacctttg ctggaaaact
     1081 tgtaaattct gctagaccct ctgtaaattc cgctagacct ttgtgtgttt tttttgttta
     1141 tattcaagtg gttataattt atagaataaa gaaagaataa aaaaagataa aaagaataga
     1201 tcccagccct gtgtataact cactacttta gtcagttccg cagtattaca aaaggatgtc
     1261 gcaaacgctg tttgctcctc tacaaaacag accttaaaac cctaaaggct taagtagcac
     1321 cctcgcaagc tcgggcaaat cgctgaatat tccttttgtc tccgaccatc aggcacctga
     1381 gtcgctgtct ttttcgtgac attcagttcg ctgcgctcac ggctctggca gtgaatgggg
     1441 gtaaatggca ctacaggcgc cttttatgga ttcatgcaag gaaactaccc ataatacaag
     1501 aaaagcccgt cacgggcttc tcagggcgtt ttatggcggg tctgctatgt ggtgctatct
     1561 gactttttgc tgttcagcag ttcctgccct ctgattttcc agtctgacca cttcggatta
     1621 tcccgtgaca ggtcattcag actggctaat gcacccagta aggcagcggt atcatcaaca
     1681 ggcttacccg tcttactgtc aagaggacat ccggtcaaat aaaacgaaag gctcagtcga
     1741 aagactgggc ctttcgtttt gctgaggaga cttagggacc ctacaacacg cacggtgtta
     1801 cattaggcac cccgggcttt acactttatg cttccggctc gtatgttgtg tcgaccgacc
     1861 tgcaggtgca gtaaggagga aaaaaaaatg aaaaaattat tattcgcaat tcctttagtt
     1921 gttcctttct attctcactc cgctgaaact gttgaaagtt gtttagcaaa accccataca
     1981 gaaaattcat ttactaacgt ctggaaagac gacaaaactt tagatcgtta cgctaactat
     2041 gagggctgtc tgtggaatgc tacaggcgtt gtagtttgta ctggtgacga aactcagtgt
     2101 tacggtacat gggttcctat tgggcttgct atccctgaaa atgagggtgg tggctctgag
     2161 ggtggcggtt ctgagggtgg cggttctgag ggtggcggta ctaaacctcc tgagtacggt
     2221 gatacaccta ttccgggcta tacttatatc aaccctctcg acggcactta tccgcctggt
     2281 actgagcaaa accccgctaa tcctaatcct tctcttgagg agtctcagcc tcttaatact
     2341 ttcatgtttc agaataatag gttccgaaat aggcaggggg cattaactgt ttatacgggc
     2401 actgttactc aaggcactga ccccgttaaa acttattacc agtacactcc tgtatcatca
     2461 aaagccatgt atgacgctta ctggaacggt aaattcagag actgcgcttt ccattctggc
     2521 tttaatgagg atccattcgt ttgtgaatat caaggccaat cgtctgacct gcctcaacct
     2581 cctgtcaatg ctggcggcgg ctctggtggt ggttctggtg gcggctctga gggtggtggc
     2641 tctgagggtg gcggttctga gggtggcggc tctgagggag gcggttccgg tggtggctct
     2701 ggttccggtg attttgatta tgaaaagatg gcaaacgcta ataagggggc tatgaccgaa
     2761 aatgccgatg aaaacgcgct acagtctgac gctaaaggca aacttgattc tgtcgctact
     2821 gattacggtg ctgctatcga tggtttcatt ggtgacgttt ccggccttgc taatggtaat
     2881 ggtgctactg gtgattttgc tggctctaat tcccaaatgg ctcaagtcgg tgacggtgat
     2941 aattcacctt taatgaataa tttccgtcaa tatttacctt ccctccctca atcggttgaa
     3001 tgtcgccctt ttgtctttgg cgctggtaaa ccttacgagt tcagtatcga ctgcgataag
     3061 atcaacctgt tccgcggtgt ctttgcgttt cttttatatg ttgccacctt tatgtatgta
     3121 ttttctacgt ttgctaacat actgcgtaat aaggagtctt aatgaaattt ggaaactttt
     3181 tgcttacata ccaacctccc caattttccc aaacagaggt aatgaaacgt ttggttaaat
     3241 taggtcgcat ctctgaggag tgtggttttg ataccgtatg gttactggag catcatttca
     3301 cggagtttgg tttgcttggt aacccttatg tcgctgctgc atatttactt ggcgcgacta
     3361 aaaaattgaa tgtaggaact gccgctattg ttcttcccac agcccatcca gtacgccaac
     3421 ttgaagatgt gaatttattg gatcaaatgt caaaaggacg atttcggttt ggtatttgcc
     3481 gagggcttta caacaaggac tttcgcgtat tcggcacaga tatgaataac agtcgcgcct
     3541 tagcggaatg ctggtacggg ctgataaaga atggcatgac agagggatat atggaagctg
     3601 ataatgaaca tatcaagttc cataaggtaa aagtaaaccc cgcggcgtat agcagaggtg
     3661 gcgcaccggt ttatgtggtg gctgaatcag cttcgacgac tgagtgggct gctcaatttg
     3721 gcctaccgat gatattaagt tggattataa atactaacga aaagaaagca caacttgagc
     3781 tttataatga agtggctcaa gaatatgggc acgatattca taatatcgac cattgcttat
     3841 catatataac atctgtagat catgactcaa ttaaagcgaa agagatttgc cggaaatttc
     3901 tggggcattg gtatgattct tatgtgaatg ctacgactat ttttgatgat tcagaccaaa
     3961 caagaggtta tgatttcaat aaagggcagt ggcgtgactt tgtattaaaa ggacataaag
     4021 atactaatcg ccgtattgat tacagttacg aaatcaatcc cgtgggaacg ccgcaggaat
     4081 gtattgacat aattcaaaaa gacattgatg ctacaggaat atcaaatatt tgttgtggat
     4141 ttgaagctaa tggaacagta gacgaaatta ttgcttccat gaagctcttc cagtctgatg
     4201 tcatgccatt tcttaaagaa aaacaacgtt cgctattata ttatggcggt ggcggtagcg
     4261 gcggtggcgg tagcggcggt ggcggtagcg gcggtggcgg tagcaaattt ggattgttct
     4321 tccttaactt catcaattca acaactgttc aagaacagag tatagttcgc atgcaggaaa
     4381 taacggagta tgttgataag ttgaattttg aacagatttt agtgtatgaa aatcattttt
     4441 cagataatgg tgttgtcggc gctcctctga ctgtttctgg ttttctgctc ggtttaacag
     4501 agaaaattaa aattggttca ttaaatcaca tcattacaac tcatcatcct gtccgcatag
     4561 cggaggaagc ttgcttattg gatcagttaa gtgaagggag atttatttta gggtttagtg
     4621 attgcgaaaa aaaagatgaa atgcattttt ttaatcgccc ggttgaatat caacagcaac
     4681 tatttgaaga gtgttatgaa atcattaacg atgctttaac aacaggctat tgtaatccag
     4741 ataacgattt ttatagcttc cctaaaatat ctgtaaatcc ccatgcttat acgccaggcg
     4801 gacctcggaa atatgtaaca gcaaccagtc atcatattgt tgagtgggcg gccaaaaaag
     4861 gtattcctct catctttaag tgggatgatt ctaatgatgt tagatatgaa tatgctgaaa
     4921 gatataaagc cgttgcggat aaatatgacg ttgacctatc agagatagac catcagttaa
     4981 tgatattagt taactataac gaagatagta ataaagctaa acaagagacg cgtgcattta
     5041 ttagtgatta tgttcttgaa atgcacccta atgaaaattt cgaaaataaa cttgaagaaa
     5101 taattgcaga aaacgctgtc ggaaattata cggagtgtat aactgcggct aagttggcaa
     5161 ttgaaaagtg tggtgcgaaa agtgtattgc tgtcctttga accaatgaat gatttgatga
     5221 gccaaaaaaa tgtaatcaat attgttgatg ataatattaa gaagtaccac acggaatata
     5281 cctaaacgga gccaatgtac gcaaaaaacc ccgcttcagc ggggtttttt cgccaaaaaa
     5341 aaccccgccc tgtcaggggc ggggtttttt tttagggccc tactgactgt tagcgtttcg
     5401 gcgccggata atgcagcgtg gtgatcagac catccgcaac cgtgctatgg tgatgcacca
     5461 gttctgccag ggtattgaag cgagattcac tagaaacgta cagtttgcca tcgctggcgg
     5521 tgttgatacg gtagtgatac acgcgacctt cataacgcag agaaatgctg cgctggcccg
     5581 ggctactttc actttcacga accagaaaag aaccattaat gccagagctc agcaggtatt
     5641 cggccgcgtt acggctcaca gggccatgat accagctgtg tttttccaga ctcaagtcct
     5701 cttcagaaat aagcttttgt tccgcggccg cacgacgacc ttcagcaata gcggtaacgg
     5761 cttgtaattc agcggcttcc tgctcttgct gttcctggcg ttcgcgaacg tcgaggatct
     5821 ggttgttgat cagaccttct tcgatttcgc gcagcgcgat tacagtggtt ttatcgtttt
     5881 cttccggtac cagcggatcc tttccgccta cctgcatctg acgagcgcga cgcgcggcga
     5941 ccagtaccag gtcaaaacgg ttaccaattt tctctacagc gtcctgaaca gttacgcgtg
     6001 ccattttttt ttcctcctta taagggggta gggggtctag atttttgtcg aactattcat
     6061 ttcacttttc tctatcactg atagggagtg gtaaaataac tctatcaatg atagagtgtc
     6121 aatctctatc actgataggg atcatctggc cattcgatgg acttaattaa cggcactgcg
     6181 aaaaaacccc gccgaagcgg ggttttttgc gaaatataat gaccctcttg ataacccaag
     6241 agggcatttt ttaatgccca tggcgtttac caatgcttaa tcagtgaggc acctatctca
     6301 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     6361 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     6421 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     6481 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     6541 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     6601 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     6661 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     6721 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     6781 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     6841 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     6901 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     6961 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     7021 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     7081 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     7141 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     7201 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     7261 gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc
     7321 ttaggccaaa taggccgt
//
