LOCUS       sequence-148357-        4640 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 4640)
  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 4640)
  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: pAB082d
FEATURES             Location/Qualifiers
     source          1..4640
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     489..508
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     742..759
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    774..914
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(840..859)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1000..1022
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1016..1207)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             1547..1819
                     /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"
     CDS             1829..1858
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(2582..3373)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(3374..3445)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(3475..4557)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     4519..4538
                     /label=LacI-R
                     /note="LacI, reverse primer"
     rep_origin      join(4640,1..588)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
       61 gcggtggttt gtttgccgga tcaagagcta ccacctcttt ttccgaaggt aactggcttc
      121 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
      181 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
      241 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
      301 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
      361 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
      421 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
      481 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
      541 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
      601 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
      661 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
      721 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
      781 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatggtg cactctcagt
      841 acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact
      901 gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
      961 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     1021 ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt
     1081 cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca
     1141 gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt
     1201 tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga
     1261 aacgagagag gatgctcacg atacgggtta ctgatgatga acaagaggac atccggtcaa
     1321 ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttgctgagga ttctagagca
     1381 cagctaacac cacgtcgtcc ctatctgctg ccctaggtct atgagtggtt gctggataac
     1441 tttacgggca tgcataaggc tcggtatcta tattcaggga gaccacaacg gtttccctct
     1501 acaaataatt ttgtttaact tttactagag taaggaggaa aaaaaaatgg cacgcgtaac
     1561 tgttcaggac gctgtagaga aaattggtaa ccgttttgac ctggtactgg tcgccgcgcg
     1621 tcgcgctcgt cagatgcagg taggcggaaa ggatccgctg gtaccggaag aaaacgataa
     1681 aaccactgta atcgcgctgc gcgaaatcga agaaggtctg atcaacaacc agatcctcga
     1741 cgttcgcgaa cgccaggaac agcaagagca ggaagccgct gaattacaag ccgttaccgc
     1801 tattgctgaa ggtcgtcgtg cggccgcgga acaaaagctt atttctgaag aggacttgga
     1861 tagtagaaga tatccaattc gaacagtttc ccaattaaca agagaaattt atacaaaccc
     1921 agtattagaa aattttgatg gtagttttcg aggctcggct cagggcatag aaagaagtat
     1981 taggagtcca catttgatgg atatacttaa cagtataacc atctatacgg atgctcatag
     2041 gggttattat tattggtcag ggcatcaaat aatggcttct cctgtagggt tttcggggcc
     2101 agaattcact tttccgctat atggaactat gggaaatgca gctccacaac aacgtattgt
     2161 tgctcaacta ggtcagggcg tgtatagaac attatcgtcc actttatata gaagaccttt
     2221 taatataggg ataaataatc aacaactatc tgttcttgac gggacagaat ttgcttatgg
     2281 aacctcctca aatttgccat ccgctgtata cagaaaaagc ggaacggtag attcgctgga
     2341 tgaaataccg ccacagaata acaacgtgcc acctaggcaa ggatttagtc atcgattaag
     2401 ccatgtttca atgtttcgtt caggctttag taatagtagt gtaagtataa taagagctcc
     2461 tatgttctct tggatacatc gtagtgctga atttaataat taacttaatt aacggcactc
     2521 ctcagcaaat ataatgaccc tcttgataac ccaagagggc attttttaat gcccatggcg
     2581 tttatttgcc gactaccttg gtgatctcgc ctttcacgta gtggacaaat tcttccaact
     2641 gatctgcgcg cgaggccaag cgatcttctt cttgtccaag ataagcctgt ctagcttcaa
     2701 gtatgacggg ctgatactgg gccggcaggc gctccattgc ccagtcggca gcgacatcct
     2761 tcggcgcgat tttgccggtt actgcgctgt accaaatgcg ggacaacgta agcactacat
     2821 ttcgctcatc gccagcccag tcgggcggcg agttccatag cgttaaggtt tcatttagcg
     2881 cctcaaatag atcctgttca ggaaccggat caaagagttc ctccgccgct ggacctacca
     2941 aggcaacgct atgttctctt gcttttgtca gcaagatagc cagatcaatg tcgatcgtgg
     3001 ctggctcgaa gatacctgca agaatgtcat tgcgctgcca ttctccaaat tgcagttcgc
     3061 gcttagctgg ataacgccac ggaatgatgt cgtcgtgcac aacaatggtg acttctacag
     3121 cgcggagaat ctcgctctct ccaggggaag ccgaagtttc caaaaggtcg ttgatcaaag
     3181 ctcgccgcgt tgtttcatca agccttacgg tcaccgtaac cagcaaatca atatcactgt
     3241 gtggcttcag gccgccatcc actgcggagc cgtacaaatg tacggccagc aacgtcggtt
     3301 cgagatggcg ctcgatgacg ccaactacct ctgatagttg agtcgatact tcggcgatca
     3361 ccgcttccct catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     3421 tcatgagcgt atacatattt gaatgtattt agaaaaatat taattgcgtt gcgctcactg
     3481 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     3541 gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg
     3601 caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct
     3661 ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aatggcggga tataacatga
     3721 gctgtcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc gcagcccgga
     3781 ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt
     3841 gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca
     3901 gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc
     3961 agccagacgc agacgcgccg agacagaact taatgggccc gctaacagtg cgatttgctg
     4021 gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat
     4081 aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca
     4141 ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact
     4201 gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc
     4261 taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac
     4321 aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg
     4381 tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc
     4441 ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac
     4501 ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt
     4561 caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgca
     4621 ccataggcca aataggccgt
//
