LOCUS       sequence-148355-        4427 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 4427)
  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 4427)
  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: pAB082b
FEATURES             Location/Qualifiers
     source          1..4427
                     /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(2369..3160)
                     /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(3161..3232)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(3262..4344)
                     /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     4306..4325
                     /label=LacI-R
                     /note="LacI, reverse primer"
     rep_origin      join(4427,1..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 aggacttggc
     1861 atcggatagt attactcaaa tccctgcagt gaagggaaac tttcttttta atggttctgt
     1921 aatttcagga ccaggattta ctggtgggga cttagttaga ttaaatagta gtggaaataa
     1981 cattcagaat agagggtata ttgaagttcc aattcacttc ccatcgacat ctaccagata
     2041 tcgagttcgt gtacggtatg cttctgtaac cccgattcac ctcaacgtta attggggtaa
     2101 ttcatccatt ttttccaata cagtaccagc tacagctacg tcattagata atctacaatc
     2161 aagtgatttt ggttattttg aaagtgccaa tgcttttaca tcttcattag gtaatatagt
     2221 aggtgttaga aattttagtg ggactgcagg agtgataata gacagatttg aatttattcc
     2281 agttacttaa cttaattaac ggcactcctc agcaaatata atgaccctct tgataaccca
     2341 agagggcatt ttttaatgcc catggcgttt atttgccgac taccttggtg atctcgcctt
     2401 tcacgtagtg gacaaattct tccaactgat ctgcgcgcga ggccaagcga tcttcttctt
     2461 gtccaagata agcctgtcta gcttcaagta tgacgggctg atactgggcc ggcaggcgct
     2521 ccattgccca gtcggcagcg acatccttcg gcgcgatttt gccggttact gcgctgtacc
     2581 aaatgcggga caacgtaagc actacatttc gctcatcgcc agcccagtcg ggcggcgagt
     2641 tccatagcgt taaggtttca tttagcgcct caaatagatc ctgttcagga accggatcaa
     2701 agagttcctc cgccgctgga cctaccaagg caacgctatg ttctcttgct tttgtcagca
     2761 agatagccag atcaatgtcg atcgtggctg gctcgaagat acctgcaaga atgtcattgc
     2821 gctgccattc tccaaattgc agttcgcgct tagctggata acgccacgga atgatgtcgt
     2881 cgtgcacaac aatggtgact tctacagcgc ggagaatctc gctctctcca ggggaagccg
     2941 aagtttccaa aaggtcgttg atcaaagctc gccgcgttgt ttcatcaagc cttacggtca
     3001 ccgtaaccag caaatcaata tcactgtgtg gcttcaggcc gccatccact gcggagccgt
     3061 acaaatgtac ggccagcaac gtcggttcga gatggcgctc gatgacgcca actacctctg
     3121 atagttgagt cgatacttcg gcgatcaccg cttccctcat actcttcctt tttcaatatt
     3181 attgaagcat ttatcagggt tattgtctca tgagcgtata catatttgaa tgtatttaga
     3241 aaaatattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     3301 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt
     3361 ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg
     3421 agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat
     3481 ggtggttaat ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga
     3541 gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat
     3601 ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt
     3661 ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg
     3721 attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa
     3781 tgggcccgct aacagtgcga tttgctggtg acccaatgcg accagatgct ccacgcccag
     3841 tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc
     3901 aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc
     3961 cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc
     4021 tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg
     4081 atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga
     4141 ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg
     4201 aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg
     4261 gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac
     4321 atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta
     4381 tcatgccata ccgcgaaagg ttttgcacca taggccaaat aggccgt
//
