LOCUS       sequence-148336-        8425 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Directed Evolution of Protein-Protein Interactions.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8425)
  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 8425)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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: pAB088h
FEATURES             Location/Qualifiers
     source          1..8425
                     /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             3184..4263
                     /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             4327..5307
                     /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(6078..6101)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      complement(6933..6960)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(7052..7138)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     7291..7308
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     CDS             complement(join(7354..8145,8146..8214))
                     /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     7977..7996
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8215..8319)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8387..8405
                     /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 ctgcttttcc agtctgacca cttcggctta
     1621 tcccgtgaca ggtcattcag actggctaat gcacccagta aggcagcggg ctcatcaaca
     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 tgaaaaaatt
     1921 attattcgca attcctttag ttgttccttt ctattctcac tccgctgaaa ctgttgaaag
     1981 ttgtttagca aaaccccata cagaaaattc atttactaac gtctggaaag acgacaaaac
     2041 tttagatcgt tacgctaact atgagggctg tctgtggaat gctacaggcg ttgtagtttg
     2101 tactggtgac gaaactcagt gttacggtac atgggttcct attgggcttg ctatccctga
     2161 aaatgagggt ggtggctctg agggtggtgg ttctgagggt ggcggttctg agggtggcgg
     2221 tactaaacct cctgagtacg gtgatacacc tattccgggc tatacttata tcaaccctct
     2281 cgacggcact tatccgcctg gtactgagca aaaccccgct aatcctaatc cttctcttga
     2341 ggagtctcag cctcttaata ctttcatgtt tcagaataat aggttccgaa ataggcaggg
     2401 ggcattaact gtttatacgg gcactgttac tcaaggcact gaccccgtta aaacttatta
     2461 ccagtacact cctgtatcat caaaagccat gtatgacgct tactggaacg gtaaattcag
     2521 agactgcgct ttccattctg gctttaatga ggatccattc gtttgtgaat atcaaggcca
     2581 atcgtctgac ctgcctcaac ctcctgtcaa tgctggcggc ggctctggtg gtggttctgg
     2641 tggcggctct gagggtggtg gctctgaggg tggcggttct gagggtggcg gctctgaggg
     2701 aggcggttcc ggtggtggct ctggttccgg tgattttgat tatgaaaaga tggcaaacgc
     2761 taataagggg gctatgaccg aaaatgccga tgaaaacgcg ctacagtctg acgctaaagg
     2821 caaacttgat tctgtcgcta ctgattacgg tgctgctatc gatggtttca ttggtgacgt
     2881 ttccggcctt gctaatggta atggtgctac tggtgatttt gctggctcta attcccaaat
     2941 ggctcaagtc ggtgacggtg ataattcacc tttaatgaat aatttccgtc aatatttacc
     3001 ttccctccct caatcggttg aatgtcgccc ttttgtcttt ggcgctggta aaccttacga
     3061 gttcagtatc gactgcgata agatcaacct gttccgcggt gtctttgcgt ttcttttata
     3121 tgttgccacc tttatgtatg tattttctac gtttgctaac atactgcgta ataaggagtc
     3181 ttaatgaaat ttggaaactt tttgcttaca taccaacctc cccaattttc ccaaacagag
     3241 gtaatgaaac gtttggttaa attaggtcgc atctctgagg agtgtggttt tgataccgta
     3301 tggttactgg agcatcattt cacggagttt ggtttgcttg gtaaccctta tgtcgctgct
     3361 gcatatttac ttggcgcgac taaaaaattg aatgtaggaa ctgccgctat tgttcttccc
     3421 acagcccatc cagtacgcca acttgaagat gtgaatttat tggatcaaat gtcaaaagga
     3481 cgatttcggt ttggtatttg ccgagggctt tacaacaagg actttcgcgt attcggcaca
     3541 gatatgaata acagtcgcgc cttagcggaa tgctggtacg ggctgataaa gaatggcatg
     3601 acagagggat atatggaagc tgataatgaa catatcaagt tccataaggt aaaagtaaac
     3661 cccgcggcgt atagcagagg tggcgcaccg gtttatgtgg tggctgaatc agcttcgacg
     3721 actgagtggg ctgctcaatt tggcctaccg atgatattaa gttggattat aaatactaac
     3781 gaaaagaaag cacaacttga gctttataat gaagtggctc aagaatatgg gcacgatatt
     3841 cataatatcg accattgctt atcatatata acatctgtag atcatgactc aattaaagcg
     3901 aaagagattt gccggaaatt tctggggcat tggtatgatt cttatgtgaa tgctacgact
     3961 atttttgatg attcagacca aacaagaggt tatgatttca ataaagggca gtggcgtgac
     4021 tttgtattaa aaggacataa agatactaat cgccgtattg attacagtta cgaaatcaat
     4081 cccgtgggaa cgccgcagga atgtattgac ataattcaaa aagacattga tgctacagga
     4141 atatcaaata tttgttgtgg atttgaagct aatggaacag tagatgaaat tattgcttcc
     4201 atgaagctct tccagtctga tgtcatgcca tttcttaaag aaaaacaacg ttcgctatta
     4261 tattatggcg gtggcggtag cggcggtggc ggtagcggcg gtggcggtag cggcggtggc
     4321 ggtagcaaat ttggattgtt cttccttaac ttcatcaatt caacaactgt tcaagaacag
     4381 agtatagttc gcatgcagga aataacggag tatgttgata agttgaattt tgaacagatt
     4441 ttagtgtatg aaaatcattt ttcagataat ggtgttgtcg gcgctcctct gactgtttct
     4501 ggttttctgc tcggtttaac agagaaaatt aaaattggtt cattaaatca catcattaca
     4561 actcatcatc ctgtccgcat agcggaggaa gcttgcttat tggatcagtt aagtgaaggg
     4621 agatttattt tagggtttag tgattgcgaa aaaaaagatg aaatgcattt ttttaatcgc
     4681 ccggttgaat atcaacagca actatttgaa gagtgttatg aaatcattaa cgatgcttta
     4741 acaacaggct attgtaatcc agataacgat ttttatagct tccctaaaat atctgtaaat
     4801 ccccatgctt atacgccagg cggacctcgg aaatatgtaa cagcaaccag tcatcatatt
     4861 gttgagtggg cggccaaaaa aggtattcct ctcatcttta agtgggatga ttctaatgat
     4921 gttagatatg aatatgctga aagatataaa gccgttgcgg ataaatatga cgttgaccta
     4981 tcagagatag accatcagtt aatgatatta gttaactata acgaagatag taataaagct
     5041 aaacaagaga cgcgtgcatt tattagtgat tatgttcttg aaatgcaccc taatgaaaat
     5101 ttcgaaaata aacttgaaga aataattgca gaaaacgctg tcggaaatta tacggagtgt
     5161 ataactgcgg ctaagttggc aattgaaaag tgtggtgcga aaagtgtatt gctgtccttt
     5221 gaaccaatga atgatttgat gagccaaaaa aatgtaatca atattgttga tgataatatt
     5281 aagaagtacc acacggaata tacctaaacg gagccaatgt acgcaaaaaa ccccgcttca
     5341 gcggggtttt ttcgccaaaa aaaaccccgc cctgtcaggg gcggggtttt tttttagggc
     5401 cctactgact gttatgtgag gctgttgtcc acgcccagct ctgggctgga gccacccaca
     5461 aagtcctcca gatgcagctc gcgagttaac agggtttcct gaatcgcacg cagcagtatt
     5521 gtgttactac ggagctgctc gatctcgtcg gtggtggcgg gcacgtggtt ccgtatgaag
     5581 tgcgcgcgca cctcggtgcg gtcgtcgcgc gcggcgccgc tgctgtccgg cgcgcgcagc
     5641 accgcgtcga tgttgcacgt caggctgaac accgacgaga acgtgtccgc gatgaaagca
     5701 cggttggagt caacgacatc cagcgtgttg ttgaacaaga agtagactct gttgagatga
     5761 gtgatcagat agatcttcac gtcagtacgg gcttctgcac cacgggtgtc tgtagccacg
     5821 acatcgaact cgaacatgcc gaccatagta tccagcgggt tcatgttgag ggacaggacg
     5881 ccgctgctcg ggtgcagcgt gaaggcggac tcacgcacgc cctccagcga ggggtccacc
     5941 accatcgtgt ccaggtccat cgtgtacgtt atgtctaggc cttcggtatg tgtcgcatga
     6001 acagtgagta gttccctgtt ggatgtatcc gtctcgtaga tgccggcggt gtagatctca
     6061 cgctcgaata ctggccgctt gtcatcgtcg tccttgtagt ctacgaattt taccctcgct
     6121 tccacgacaa caccgataat cttgcagttt ccgttgatag gagtcatagg ccatgaagga
     6181 ttcaggcctt tcaggtactt ctgaccgcca tctatgacca gtttcttgaa tgttgcttcg
     6241 ttcgcgtcag tcagtttggc tacaacaagg cttccattca ctggctcccg tccagtatct
     6301 actaacacca tatgaccttc agggatgctt tgacctacag gtgaggtcat ggaatcacct
     6361 tcaaccttca gccagaatcc attgcctaat aagttaacgt cactgtcata ccattcatca
     6421 atgtccttga tatcgtaggg ttcacaagct tcacaccacg aacgagctct aaccatgcta
     6481 atcaatggat atttcccttt gggctcaacg tgcccaacaa atctaacatt cgaatcagag
     6541 gtgccattga gcagccagtc aacacttacg ccaagagctg acgcaagttc tggtaaaaag
     6601 cgtggtcgct tagttttacc gttttcgagc tgctctatag actgctgggt agtccccacc
     6661 ttttgagcaa gttcagcctg gttaagtcca agctgaattc ttttgctttt taccctgctg
     6721 ctgatactca tttttttttc ctccttactc tagtaaaagt taaacaaaat tatttgtaga
     6781 gggaaaccgt tgtggtctcc ctgaatatag ataccgagcc ttatgcatgc ccgtaaagtt
     6841 atccagcaac cactcataga cctagggcag cagataggga cgacgtggtg ttagctgtgc
     6901 tctagaatcc tcagcgagtt tgtagaaacg caaaaaggcc atccgtcagg atggccttct
     6961 gcttaatttg atgcctggca gtttatggcg ggcgtcctgc ccgccaccct ccgggccgtt
     7021 gcttcgcaac gttcaaatcc gctcccggcg gatttgtcct actcaggaga gcgttcaccg
     7081 acaaacaaca gataaaacga aaggcccagt ctttcgactg agcctttcgt tttatttgat
     7141 gcctggcagt tccctactct cgcatgggga gaccccacac taccatcggc gctacggcgt
     7201 ttcacttctg agttcggcat ggggtcaggt gggaccaccg cgctactgcc gccaggcaaa
     7261 ttctgtttta tcagaccgct tctgcgttct gatttaatct gtatcaggct gaaaatcttc
     7321 tctcatccgc caaaacagcc atgcccatgg cgtttaccaa tgcttaatca gtgaggcacc
     7381 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     7441 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     7501 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     7561 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     7621 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     7681 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     7741 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     7801 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     7861 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     7921 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     7981 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     8041 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     8101 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     8161 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     8221 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     8281 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     8341 acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac
     8401 gaggccctta ggccaaatag gccgt
//
