LOCUS       sequence-148327-        6788 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 6788)
  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 6788)
  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: pAB078d8
FEATURES             Location/Qualifiers
     source          1..6788
                     /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"
     CDS             1910..2989
                     /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             3053..4033
                     /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(4441..4464)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      complement(5296..5323)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(5415..5501)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     5654..5671
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     CDS             complement(join(5717..6508,6509..6577))
                     /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     6340..6359
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6578..6682)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6750..6768
                     /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 taattttaat 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 tcgtacggtc aagaggacat ccgggcaaat gcaacgaaag gctcagtcga
     1741 aagactgggc ctttcgttgc gctgcggaga cttagggacc ctaaaacaag atacattgta
     1801 tgaaaataca agaaagtttg ttcattaggc accccgggct ttactcgtaa agcttccggc
     1861 gcgtatgttg tgtcgaccga cctgcaggtg cagtaaggag gaaaaaaaaa tgaaatttgg
     1921 aaactttttg cttacatacc aacctcccca attttcccaa acagaggtaa tgaaacgttt
     1981 ggttaaatta ggtcgcatct ctgaggagtg tggttttgat accgtatggt tactggagca
     2041 tcatttcacg gagtttggtt tgcttggtaa cccttatgtc gctgctgcat atttacttgg
     2101 cgcgactaaa aaattgaatg taggaactgc cgctattgtt cttcccacag cccatccagt
     2161 acgccaactt gaagatgtga atttattgga tcaaatgtca aaaggacgat ttcggtttgg
     2221 tatttgccga gggctttaca acaaggactt tcgcgtattc ggcacagata tgaataacag
     2281 tcgcgcctta gcggaatgct ggtacgggct gataaagaat ggcatgacag agggatatat
     2341 ggaagctgat aatgaacata tcaagttcca taaggtaaaa gtaaaccccg cggcgtatag
     2401 cagaggtggc gcaccggttt atgtggtggc tgaatcagct tcgacgactg agtgggctgc
     2461 tcaatttggc ctaccgatga tattaagttg gattataaat actaacgaaa agaaagcaca
     2521 acttgagctt tataatgaag tggctcaaga atatgggcac gatattcata atatcgacca
     2581 ttgcttatca tatataacat ctgtagatca tgactcaatt aaagcgaaag agatttgccg
     2641 gaaatttctg gggcattggt atgattctta tgtgaatgct acgactattt ttgatgattc
     2701 agaccaaaca agaggttatg atttcaataa agggcagtgg cgtgactttg tattaaaagg
     2761 acataaagat actaatcgcc gtattgatta cagttacgaa atcaatcccg tgggaacgcc
     2821 gcaggaatgt attgacataa ttcaaaaaga cattgatgct acaggaatat caaatatttg
     2881 ttgtggattt gaagctaatg gaacagtaga cgaaattatt gcttccatga agctcttcca
     2941 gtctgatgtc atgccatttc ttaaagaaaa acaacgttcg ctattatatt atggcggtgg
     3001 cggtagcggc ggtggcggta gcggcggtgg cggtagcggc ggtggcggta gcaaatttgg
     3061 attgttcttc cttaacttca tcaattcaac aactgttcaa gaacagagta tagttcgcat
     3121 gcaggaaata acggagtatg ttgataagtt gaattttgaa cagattttag tgtatgaaaa
     3181 tcatttttca gataatggtg ttgtcggcgc tcctctgact gtttctggtt ttctgctcgg
     3241 tttaacagag aaaattaaaa ttggttcatt aaatcacatc attacaactc atcatcctgt
     3301 ccgcatagcg gaggaagctt gcttattgga tcagttaagt gaagggagat ttattttagg
     3361 gtttagtgat tgcgaaaaaa aagatgaaat gcattttttt aatcgcccgg ttgaatatca
     3421 acagcaacta tttgaagagt gttatgaaat cattaacgat gctttaacaa caggctattg
     3481 taatccagat aacgattttt atagcttccc taaaatatct gtaaatcccc atgcttatac
     3541 gccaggcgga cctcggaaat atgtaacagc aaccagtcat catattgttg agtgggcggc
     3601 caaaaaaggt attcctctca tctttaagtg ggatgattct aatgatgtta gatatgaata
     3661 tgctgaaaga tataaagccg ttgcggataa atatgacgtt gacctatcag agatagacca
     3721 tcagttaatg atattagtta actataacga agatagtaat aaagctaaac aagagacgcg
     3781 tgcatttatt agtgattatg ttcttgaaat gcaccctaat gaaaatttcg aaaataaact
     3841 tgaagaaata attgcagaaa acgctgtcgg aaattatacg gagtgtataa ctgcggctaa
     3901 gttggcaatt gaaaagtgtg gtgcgaaaag tgtattgctg tcctttgaac caatgaatga
     3961 tttgatgagc caaaaaaatg taatcaatat tgttgatgat aatattaaga agtaccacac
     4021 ggaatatacc taaacggagc caatgtacgc aaaaaacccc gcttcagcgg ggttttttcg
     4081 ccaaaaaaaa ccccgccctg tcaggggcgg ggtttttttt tagggcccta ctgactgtta
     4141 gcgtttcggc gccggataat gcagcgtggt gatcagacca tccgcaaccg tgctatggtg
     4201 atgcaccagt tctgccaggg tattgaagcg agattcacta gaaacgtaca gtttgccatc
     4261 gctggcggtg ttgatacggt agtgatacac gcgaccttca taacgcagag aaatgctgcg
     4321 ctggcccggg ctactttcac tttcacgaac cagaaaagaa ccattaatgc cagagctcag
     4381 caggtattcg gccgcgttac ggctcacagg gccatgatac cagctgtgtt tttccagact
     4441 cttgtcatcg tcgtccttgt agtctacgaa ttttaccctc gcttccacga caacaccgat
     4501 aatcttgcag tttccgttga taggagtcat aggccatgaa ggattcaggc ctttcaggta
     4561 cttctgaccg ccatctatga ccagtttctt gaatgttgct tcgttcgcgt cagtcagttt
     4621 ggctacaaca aggcttccat tcactggctc ccgtccagta tctactaaca ccatatgacc
     4681 ttcagggatg ctttgaccta caggtgaggt catggaatca ccttcaacct tcagccagaa
     4741 tccattgcct aataagttaa cgtcactgtc ataccattca tcaatgtcct tgatatcgta
     4801 gggttcacaa gcttcacacc acgaacgagc tctaaccatg ctaatcaatg gatatttccc
     4861 tttgggctca acgtgcccaa caaatctaac attcgaatca gaggtgccat tgagcagcca
     4921 gtcaacactt acgccaagag ctgacgcaag ttctggtaaa aagcgtggtc gcttagtttt
     4981 accgttttcg agctgctcta tagactgctg ggtagtcccc accttttgag caagttcagc
     5041 ctggttaagt ccaagctgaa ttcttttgct ttttaccctg ctgctgatac tcattttttt
     5101 ttcctcctta ctctagtaaa agttaaacaa aattatttgt agagggaaac cgttgtggtc
     5161 tccctgaata tagataccga gccttatgca tgcccgtaaa gttatccagc aaccactcat
     5221 agacctaggg cagcagatag ggacgacgtg gtgttagctg tgctctagaa tcctcagcga
     5281 gtttgtagaa acgcaaaaag gccatccgtc aggatggcct tctgcttaat ttgatgcctg
     5341 gcagtttatg gcgggcgtcc tgcccgccac cctccgggcc gttgcttcgc aacgttcaaa
     5401 tccgctcccg gcggatttgt cctactcagg agagcgttca ccgacaaaca acagataaaa
     5461 cgaaaggccc agtctttcga ctgagccttt cgttttattt gatgcctggc agttccctac
     5521 tctcgcatgg ggagacccca cactaccatc ggcgctacgg cgtttcactt ctgagttcgg
     5581 catggggtca ggtgggacca ccgcgctact gccgccaggc aaattctgtt ttatcagacc
     5641 gcttctgcgt tctgatttaa tctgtatcag gctgaaaatc ttctctcatc cgccaaaaca
     5701 gccatgccca tggcgtttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5761 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5821 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5881 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5941 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6001 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6061 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6121 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6181 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6241 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6301 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     6361 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6421 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6481 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6541 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6601 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6661 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     6721 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc ttaggccaaa
     6781 taggccgt
//
