LOCUS       sequence-148360-        5075 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 5075)
  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 5075)
  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: pAB082g
FEATURES             Location/Qualifiers
     source          1..5075
                     /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(3017..3808)
                     /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(3809..3880)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(3910..4992)
                     /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     4954..4973
                     /label=LacI-R
                     /note="LacI, reverse primer"
     rep_origin      join(5075,1..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 ataattgcat cggatagtat
     2521 tactcaaatc cctgcagtga agggaaactt tctttttaat ggttctgtaa tttcaggacc
     2581 aggatttact ggtggggact tagttagatt aaatagtagt ggaaataaca ttcagaatag
     2641 agggtatatt gaagttccaa ttcacttccc atcgacatct accagatatc gagttcgtgt
     2701 acggtatgct tctgtaaccc cgattcacct caacgttaat tggggtaatt catccatttt
     2761 ttccaataca gtaccagcta cagctacgtc attagataat ctacaatcaa gtgattttgg
     2821 ttattttgaa agtgccaatg cttttacatc ttcattaggt aatatagtag gtgttagaaa
     2881 ttttagtggg actgcaggag tgataataga cagatttgaa tttattccag ttacttaact
     2941 taattaacgg cactcctcag caaatataat gaccctcttg ataacccaag agggcatttt
     3001 ttaatgccca tggcgtttat ttgccgacta ccttggtgat ctcgcctttc acgtagtgga
     3061 caaattcttc caactgatct gcgcgcgagg ccaagcgatc ttcttcttgt ccaagataag
     3121 cctgtctagc ttcaagtatg acgggctgat actgggccgg caggcgctcc attgcccagt
     3181 cggcagcgac atccttcggc gcgattttgc cggttactgc gctgtaccaa atgcgggaca
     3241 acgtaagcac tacatttcgc tcatcgccag cccagtcggg cggcgagttc catagcgtta
     3301 aggtttcatt tagcgcctca aatagatcct gttcaggaac cggatcaaag agttcctccg
     3361 ccgctggacc taccaaggca acgctatgtt ctcttgcttt tgtcagcaag atagccagat
     3421 caatgtcgat cgtggctggc tcgaagatac ctgcaagaat gtcattgcgc tgccattctc
     3481 caaattgcag ttcgcgctta gctggataac gccacggaat gatgtcgtcg tgcacaacaa
     3541 tggtgacttc tacagcgcgg agaatctcgc tctctccagg ggaagccgaa gtttccaaaa
     3601 ggtcgttgat caaagctcgc cgcgttgttt catcaagcct tacggtcacc gtaaccagca
     3661 aatcaatatc actgtgtggc ttcaggccgc catccactgc ggagccgtac aaatgtacgg
     3721 ccagcaacgt cggttcgaga tggcgctcga tgacgccaac tacctctgat agttgagtcg
     3781 atacttcggc gatcaccgct tccctcatac tcttcctttt tcaatattat tgaagcattt
     3841 atcagggtta ttgtctcatg agcgtataca tatttgaatg tatttagaaa aatattaatt
     3901 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     3961 atcggccaac gcgcggggag aggcggtttg cgtattgggc gccagggtgg tttttctttt
     4021 caccagtgag acgggcaaca gctgattgcc cttcaccgcc tggccctgag agagttgcag
     4081 caagcggtcc acgctggttt gccccagcag gcgaaaatcc tgtttgatgg tggttaatgg
     4141 cgggatataa catgagctgt cttcggtatc gtcgtatccc actaccgaga tatccgcacc
     4201 aacgcgcagc ccggactcgg taatggcgcg cattgcgccc agcgccatct gatcgttggc
     4261 aaccagcatc gcagtgggaa cgatgccctc attcagcatt tgcatggttt gttgaaaacc
     4321 ggacatggca ctccagtcgc cttcccgttc cgctatcggc tgaatttgat tgcgagtgag
     4381 atatttatgc cagccagcca gacgcagacg cgccgagaca gaacttaatg ggcccgctaa
     4441 cagtgcgatt tgctggtgac ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc
     4501 ttcatgggag aaaataatac tgttgatggg tgtctggtca gagacatcaa gaaataacgc
     4561 cggaacatta gtgcaggcag cttccacagc aatggcatcc tggtcatcca gcggatagtt
     4621 aatgatcagc ccactgacgc gttgcgcgag aagattgtgc accgccgctt tacaggcttc
     4681 gacgccgctt cgttctacca tcgacaccac cacgctggca cccagttgat cggcgcgaga
     4741 tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc agactggagg tggcaacgcc
     4801 aatcagcaac gactgtttgc ccgccagttg ttgtgccacg cggttgggaa tgtaattcag
     4861 ctccgccatc gccgcttcca ctttttcccg cgttttcgca gaaacgtggc tggcctggtt
     4921 caccacgcgg gaaacggtct gataagagac accggcatac tctgcgacat cgtataacgt
     4981 tactggtttc acattcacca ccctgaattg actctcttcc gggcgctatc atgccatacc
     5041 gcgaaaggtt ttgcaccata ggccaaatag gccgt
//
