LOCUS       sequence-148354-        5822 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 5822)
  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 5822)
  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: pAB082a
FEATURES             Location/Qualifiers
     source          1..5822
                     /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(3764..4555)
                     /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(4556..4627)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(4657..5739)
                     /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     5701..5720
                     /label=LacI-R
                     /note="LacI, reverse primer"
     rep_origin      join(5822,1..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 taacaatccg aacatcaatg aatgcattcc ttataattgt ttaagtaacc ctgaagtaga
     1921 agtattaggt ggagaaagaa tagaaactgg ttacacccca atcgatattt ccttgtcgct
     1981 aacgcaattt cttttgagtg aatttgttcc cggtgctgga tttgtgttag gactagttga
     2041 tataatatgg ggaatttttg gtccctctca atgggacgca tttcttgtac aaattgaaca
     2101 gttaattaac caaagaatag aagaattcgc taggaaccaa gccatttcta gattagaagg
     2161 actaagcaat ctttatcaaa tttacgcaga atcttttaga gagtgggaag cagatcctac
     2221 taatccagca ttaagagaag agatgcgtat tcaattcaat gacatgaaca gtgcccttac
     2281 aaccgctatt cctctttttg cagttcaaaa ttatcaagtt cctcttttat cagtatatgt
     2341 tcaagctgca aatttacatt tatcagtttt gagagatgtt tcagtgtttg gacaaaggtg
     2401 gggatttgat gccgcgacta tcaatagtcg ttataatgat ttaactaggc ttattggcaa
     2461 ctatacagat tatgctgtac gctggtacaa tacgggatta gaacgtgtat ggggaccgga
     2521 ttctagagat tgggtaaggt ataatcaatt tagaagagaa ttaacactaa ctgtattaga
     2581 tatcgttgct ctgttcccga attatgatag tagaagatat ccaattcgaa cagtttccca
     2641 attaacaaga gaaatttata caaacccagt attagaaaat tttgatggta gttttcgagg
     2701 ctcggctcag ggcatagaaa gaagtattag gagtccacat ttgatggata tacttaacag
     2761 tataaccatc tatacggatg ctcatagggg ttattattat tggtcagggc atcaaataat
     2821 ggcttctcct gtagggtttt cggggccaga attcactttt ccgctatatg gaactatggg
     2881 aaatgcagct ccacaacaac gtattgttgc tcaactaggt cagggcgtgt atagaacatt
     2941 atcgtccact ttatatagaa gaccttttaa tatagggata aataatcaac aactatctgt
     3001 tcttgacggg acagaatttg cttatggaac ctcctcaaat ttgccatccg ctgtatacag
     3061 aaaaagcgga acggtagatt cgctggatga aataccgcca cagaataaca acgtgccacc
     3121 taggcaagga tttagtcatc gattaagcca tgtttcaatg tttcgttcag gctttagtaa
     3181 tagtagtgta agtataataa gagctcctat gttctcttgg atacatcgta gtgctgaatt
     3241 taataatata attgcatcgg atagtattac tcaaatccct gcagtgaagg gaaactttct
     3301 ttttaatggt tctgtaattt caggaccagg atttactggt ggggacttag ttagattaaa
     3361 tagtagtgga aataacattc agaatagagg gtatattgaa gttccaattc acttcccatc
     3421 gacatctacc agatatcgag ttcgtgtacg gtatgcttct gtaaccccga ttcacctcaa
     3481 cgttaattgg ggtaattcat ccattttttc caatacagta ccagctacag ctacgtcatt
     3541 agataatcta caatcaagtg attttggtta ttttgaaagt gccaatgctt ttacatcttc
     3601 attaggtaat atagtaggtg ttagaaattt tagtgggact gcaggagtga taatagacag
     3661 atttgaattt attccagtta cttaacttaa ttaacggcac tcctcagcaa atataatgac
     3721 cctcttgata acccaagagg gcatttttta atgcccatgg cgtttatttg ccgactacct
     3781 tggtgatctc gcctttcacg tagtggacaa attcttccaa ctgatctgcg cgcgaggcca
     3841 agcgatcttc ttcttgtcca agataagcct gtctagcttc aagtatgacg ggctgatact
     3901 gggccggcag gcgctccatt gcccagtcgg cagcgacatc cttcggcgcg attttgccgg
     3961 ttactgcgct gtaccaaatg cgggacaacg taagcactac atttcgctca tcgccagccc
     4021 agtcgggcgg cgagttccat agcgttaagg tttcatttag cgcctcaaat agatcctgtt
     4081 caggaaccgg atcaaagagt tcctccgccg ctggacctac caaggcaacg ctatgttctc
     4141 ttgcttttgt cagcaagata gccagatcaa tgtcgatcgt ggctggctcg aagatacctg
     4201 caagaatgtc attgcgctgc cattctccaa attgcagttc gcgcttagct ggataacgcc
     4261 acggaatgat gtcgtcgtgc acaacaatgg tgacttctac agcgcggaga atctcgctct
     4321 ctccagggga agccgaagtt tccaaaaggt cgttgatcaa agctcgccgc gttgtttcat
     4381 caagccttac ggtcaccgta accagcaaat caatatcact gtgtggcttc aggccgccat
     4441 ccactgcgga gccgtacaaa tgtacggcca gcaacgtcgg ttcgagatgg cgctcgatga
     4501 cgccaactac ctctgatagt tgagtcgata cttcggcgat caccgcttcc ctcatactct
     4561 tcctttttca atattattga agcatttatc agggttattg tttcatgagc gtatacatat
     4621 ttgaatgtat ttagaaaaat attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4681 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4741 attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt
     4801 caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg
     4861 aaaatcctgt ttgatggtgg ttaatggcgg gatataacat gagctgtctt cggtatcgtc
     4921 gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat
     4981 tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt
     5041 cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc
     5101 tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc
     5161 cgagacagaa cttaatgggc ccgctaacag tgcgatttgc tggtgaccca atgcgaccag
     5221 atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt
     5281 ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat
     5341 ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag
     5401 attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac
     5461 gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg
     5521 cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg
     5581 tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt
     5641 tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc
     5701 ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact
     5761 ctcttccggg cgctatcatg ccataccgcg aaaggttttg caccataggc caaataggcc
     5821 gt
//
