LOCUS       sequence-148328-        6811 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 6811)
  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 6811)
  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: pAB078d9
FEATURES             Location/Qualifiers
     source          1..6811
                     /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             1933..3012
                     /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             3076..4056
                     /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(4464..4487)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      complement(5319..5346)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(5438..5524)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     5677..5694
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     CDS             complement(join(5740..6531,6532..6600))
                     /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     6363..6382
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6601..6705)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6773..6791
                     /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 ctaacacaag aaaaactgta
     1801 tttgacaaac aagatacatt gtatgaaaat acaagaaagt ttgttcatta ggcaccccgg
     1861 gctttactcg taaagcttcc ggcgcgtatg ttgtgtcgac cgacctgcag gtgcagtaag
     1921 gaggaaaaaa aaatgaaatt tggaaacttt ttgcttacat accaacctcc ccaattttcc
     1981 caaacagagg taatgaaacg tttggttaaa ttaggtcgca tctctgagga gtgtggtttt
     2041 gataccgtat ggttactgga gcatcatttc acggagtttg gtttgcttgg taacccttat
     2101 gtcgctgctg catatttact tggcgcgact aaaaaattga atgtaggaac tgccgctatt
     2161 gttcttccca cagcccatcc agtacgccaa cttgaagatg tgaatttatt ggatcaaatg
     2221 tcaaaaggac gatttcggtt tggtatttgc cgagggcttt acaacaagga ctttcgcgta
     2281 ttcggcacag atatgaataa cagtcgcgcc ttagcggaat gctggtacgg gctgataaag
     2341 aatggcatga cagagggata tatggaagct gataatgaac atatcaagtt ccataaggta
     2401 aaagtaaacc ccgcggcgta tagcagaggt ggcgcaccgg tttatgtggt ggctgaatca
     2461 gcttcgacga ctgagtgggc tgctcaattt ggcctaccga tgatattaag ttggattata
     2521 aatactaacg aaaagaaagc acaacttgag ctttataatg aagtggctca agaatatggg
     2581 cacgatattc ataatatcga ccattgctta tcatatataa catctgtaga tcatgactca
     2641 attaaagcga aagagatttg ccggaaattt ctggggcatt ggtatgattc ttatgtgaat
     2701 gctacgacta tttttgatga ttcagaccaa acaagaggtt atgatttcaa taaagggcag
     2761 tggcgtgact ttgtattaaa aggacataaa gatactaatc gccgtattga ttacagttac
     2821 gaaatcaatc ccgtgggaac gccgcaggaa tgtattgaca taattcaaaa agacattgat
     2881 gctacaggaa tatcaaatat ttgttgtgga tttgaagcta atggaacagt agacgaaatt
     2941 attgcttcca tgaagctctt ccagtctgat gtcatgccat ttcttaaaga aaaacaacgt
     3001 tcgctattat attatggcgg tggcggtagc ggcggtggcg gtagcggcgg tggcggtagc
     3061 ggcggtggcg gtagcaaatt tggattgttc ttccttaact tcatcaattc aacaactgtt
     3121 caagaacaga gtatagttcg catgcaggaa ataacggagt atgttgataa gttgaatttt
     3181 gaacagattt tagtgtatga aaatcatttt tcagataatg gtgttgtcgg cgctcctctg
     3241 actgtttctg gttttctgct cggtttaaca gagaaaatta aaattggttc attaaatcac
     3301 atcattacaa ctcatcatcc tgtccgcata gcggaggaag cttgcttatt ggatcagtta
     3361 agtgaaggga gatttatttt agggtttagt gattgcgaaa aaaaagatga aatgcatttt
     3421 tttaatcgcc cggttgaata tcaacagcaa ctatttgaag agtgttatga aatcattaac
     3481 gatgctttaa caacaggcta ttgtaatcca gataacgatt tttatagctt ccctaaaata
     3541 tctgtaaatc cccatgctta tacgccaggc ggacctcgga aatatgtaac agcaaccagt
     3601 catcatattg ttgagtgggc ggccaaaaaa ggtattcctc tcatctttaa gtgggatgat
     3661 tctaatgatg ttagatatga atatgctgaa agatataaag ccgttgcgga taaatatgac
     3721 gttgacctat cagagataga ccatcagtta atgatattag ttaactataa cgaagatagt
     3781 aataaagcta aacaagagac gcgtgcattt attagtgatt atgttcttga aatgcaccct
     3841 aatgaaaatt tcgaaaataa acttgaagaa ataattgcag aaaacgctgt cggaaattat
     3901 acggagtgta taactgcggc taagttggca attgaaaagt gtggtgcgaa aagtgtattg
     3961 ctgtcctttg aaccaatgaa tgatttgatg agccaaaaaa atgtaatcaa tattgttgat
     4021 gataatatta agaagtacca cacggaatat acctaaacgg agccaatgta cgcaaaaaac
     4081 cccgcttcag cggggttttt tcgccaaaaa aaaccccgcc ctgtcagggg cggggttttt
     4141 ttttagggcc ctactgactg ttagcgtttc ggcgccggat aatgcagcgt ggtgatcaga
     4201 ccatccgcaa ccgtgctatg gtgatgcacc agttctgcca gggtattgaa gcgagattca
     4261 ctagaaacgt acagtttgcc atcgctggcg gtgttgatac ggtagtgata cacgcgacct
     4321 tcataacgca gagaaatgct gcgctggccc gggctacttt cactttcacg aaccagaaaa
     4381 gaaccattaa tgccagagct cagcaggtat tcggccgcgt tacggctcac agggccatga
     4441 taccagctgt gtttttccag actcttgtca tcgtcgtcct tgtagtctac gaattttacc
     4501 ctcgcttcca cgacaacacc gataatcttg cagtttccgt tgataggagt cataggccat
     4561 gaaggattca ggcctttcag gtacttctga ccgccatcta tgaccagttt cttgaatgtt
     4621 gcttcgttcg cgtcagtcag tttggctaca acaaggcttc cattcactgg ctcccgtcca
     4681 gtatctacta acaccatatg accttcaggg atgctttgac ctacaggtga ggtcatggaa
     4741 tcaccttcaa ccttcagcca gaatccattg cctaataagt taacgtcact gtcataccat
     4801 tcatcaatgt ccttgatatc gtagggttca caagcttcac accacgaacg agctctaacc
     4861 atgctaatca atggatattt ccctttgggc tcaacgtgcc caacaaatct aacattcgaa
     4921 tcagaggtgc cattgagcag ccagtcaaca cttacgccaa gagctgacgc aagttctggt
     4981 aaaaagcgtg gtcgcttagt tttaccgttt tcgagctgct ctatagactg ctgggtagtc
     5041 cccacctttt gagcaagttc agcctggtta agtccaagct gaattctttt gctttttacc
     5101 ctgctgctga tactcatttt tttttcctcc ttactctagt aaaagttaaa caaaattatt
     5161 tgtagaggga aaccgttgtg gtctccctga atatagatac cgagccttat gcatgcccgt
     5221 aaagttatcc agcaaccact catagaccta gggcagcaga tagggacgac gtggtgttag
     5281 ctgtgctcta gaatcctcag cgagtttgta gaaacgcaaa aaggccatcc gtcaggatgg
     5341 ccttctgctt aatttgatgc ctggcagttt atggcgggcg tcctgcccgc caccctccgg
     5401 gccgttgctt cgcaacgttc aaatccgctc ccggcggatt tgtcctactc aggagagcgt
     5461 tcaccgacaa acaacagata aaacgaaagg cccagtcttt cgactgagcc tttcgtttta
     5521 tttgatgcct ggcagttccc tactctcgca tggggagacc ccacactacc atcggcgcta
     5581 cggcgtttca cttctgagtt cggcatgggg tcaggtggga ccaccgcgct actgccgcca
     5641 ggcaaattct gttttatcag accgcttctg cgttctgatt taatctgtat caggctgaaa
     5701 atcttctctc atccgccaaa acagccatgc ccatggcgtt taccaatgct taatcagtga
     5761 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     5821 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     5881 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     5941 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6001 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6061 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6121 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     6181 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     6241 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     6301 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     6361 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     6421 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     6481 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     6541 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     6601 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     6661 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     6721 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     6781 tatcacgagg cccttaggcc aaataggccg t
//
