LOCUS       sequence-148346-        4309 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 4309)
  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 4309)
  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: pAB049b
FEATURES             Location/Qualifiers
     source          1..4309
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    775..915
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(841..860)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1001..1023
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1017..1208)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     protein_bind    1426..1447
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1462..1467,1468..1485,1486..1492)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1500..1516
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by 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)."
     CDS             1870..1899
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(2251..3042)
                     /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(3043..3114)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(3144..4226)
                     /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     4188..4207
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accacctctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
      781 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatatggt gcactctcag
      841 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
      901 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
      961 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     1021 aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg
     1081 tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc
     1141 agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt
     1201 ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg
     1261 aaacgagaga ggatgctcac gatacgggtt actgatgatg aacaagagga catccggtca
     1321 aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttgctgagg agacttaggg
     1381 accctggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
     1441 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
     1501 tgtgagcgga taacaatttc acacacctgc aggtgcagta aggaggaaaa aaaaatgggc
     1561 agctctgtga gtagcgttcc gaccaaactg gaagtggttg cagcaacccc gacgagcctg
     1621 ctgatttctt gggatgcccc gatgtctagt agctctgtgt attactatcg tatcacctac
     1681 ggtgaaacgg gcggtaacag cccggtgcag gaatttacgg ttccgtatag tagctctacc
     1741 gcgacgatta gtggcctgag cccgggtgtg gattacacca tcacggttta tgcatggggc
     1801 gaagatagcg cgggttacat gttcatgtat tctccgatta gtatcaatta ccgcacctgc
     1861 gcggccgcgg aacaaaagct tatttctgaa gaggacttga tgaataaaaa cattgataca
     1921 gttcgtgaaa ttattactgt tgcatctatt ttgattaaat tttccagaga agatattgtt
     1981 gagaatcgtg ctaattttat tgcatttcta aatgagattg gagtaacgca tgaaggtaga
     2041 aagttaaatc aaaattcatt ccgtaaaatt gtttctgaat taactcaaga agataagaaa
     2101 accctcatcg acgaattcaa cgagggtttt gagggtgtat atcgatatct agagatgtat
     2161 acgaacaaat aacttaatta acggcactcc tcagcaaata taatgaccct cttgataacc
     2221 caagagggca ttttttaatg cccatggcgt ttatttgccg actaccttgg tgatctcgcc
     2281 tttcacgtag tggacaaatt cttccaactg atctgcgcgc gaggccaagc gatcttcttc
     2341 ttgtccaaga taagcctgtc tagcttcaag tatgacgggc tgatactggg ccggcaggcg
     2401 ctccattgcc cagtcggcag cgacatcctt cggcgcgatt ttgccggtta ctgcgctgta
     2461 ccaaatgcgg gacaacgtaa gcactacatt tcgctcatcg ccagcccagt cgggcggcga
     2521 gttccatagc gttaaggttt catttagcgc ctcaaataga tcctgttcag gaaccggatc
     2581 aaagagttcc tccgccgctg gacctaccaa ggcaacgcta tgttctcttg cttttgtcag
     2641 caagatagcc agatcaatgt cgatcgtggc tggctcgaag atacctgcaa gaatgtcatt
     2701 gcgctgccat tctccaaatt gcagttcgcg cttagctgga taacgccacg gaatgatgtc
     2761 gtcgtgcaca acaatggtga cttctacagc gcggagaatc tcgctctctc caggggaagc
     2821 cgaagtttcc aaaaggtcgt tgatcaaagc tcgccgcgtt gtttcatcaa gccttacggt
     2881 caccgtaacc agcaaatcaa tatcactgtg tggcttcagg ccgccatcca ctgcggagcc
     2941 gtacaaatgt acggccagca acgtcggttc gagatggcgc tcgatgacgc caactacctc
     3001 tgatagttga gtcgatactt cggcgatcac cgcttccctc atactcttcc tttttcaata
     3061 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     3121 gaaaaatatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     3181 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg
     3241 gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc
     3301 tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg
     3361 atggtggtta acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc
     3421 gagatatccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc
     3481 atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg
     3541 gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt
     3601 tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt
     3661 aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc
     3721 agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca
     3781 tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca
     3841 tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc
     3901 gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt
     3961 tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg
     4021 gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg
     4081 ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg
     4141 tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg
     4201 acatcgtata acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc
     4261 tatcatgcca taccgcgaaa ggttttgcac cataggccaa ataggccgt
//
