LOCUS Exported 6788 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Directed Evolution of Protein-Protein Interactions. ACCESSION . VERSION . KEYWORDS pAB078d8 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6788) 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 6788) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6788 /organism="synthetic DNA construct" /mol_type="other DNA" CDS complement(1..951) /codon_start=1 /gene="rep101" /product="RepA protein needed for replication with the pSC101 origin" /label=Rep101 /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI" rep_origin 999..1221 /label=pSC101 ori /note="low-copy replication origin that requires the Rep101 protein" CDS 1910..2989 /codon_start=1 /gene="luxA" /product="LuxA luciferase subunit" /label=LuxA /note="from the Photorhabdus luminescens bioluminescence operon" /translation="MKFGNFLLTYQPPQFSQTEVMKRLVKLGRISEECGFDTVWLLEHH FTEFGLLGNPYVAAAYLLGATKKLNVGTAAIVLPTAHPVRQLEDVNLLDQMSKGRFRFG ICRGLYNKDFRVFGTDMNNSRALAECWYGLIKNGMTEGYMEADNEHIKFHKVKVNPAAY SRGGAPVYVVAESASTTEWAAQFGLPMILSWIINTNEKKAQLELYNEVAQEYGHDIHNI DHCLSYITSVDHDSIKAKEICRKFLGHWYDSYVNATTIFDDSDQTRGYDFNKGQWRDFV LKGHKDTNRRIDYSYEINPVGTPQECIDIIQKDIDATGISNICCGFEANGTVDEIIASM KLFQSDVMPFLKEKQRSLLY" CDS 3053..4033 /codon_start=1 /gene="luxB" /product="LuxB luciferase subunit" /label=LuxB /note="from the Photorhabdus luminescens bioluminescence operon" /translation="KFGLFFLNFINSTTVQEQSIVRMQEITEYVDKLNFEQILVYENHF SDNGVVGAPLTVSGFLLGLTEKIKIGSLNHIITTHHPVRIAEEACLLDQLSEGRFILGF SDCEKKDEMHFFNRPVEYQQQLFEECYEIINDALTTGYCNPDNDFYSFPKISVNPHAYT PGGPRKYVTATSHHIVEWAAKKGIPLIFKWDDSNDVRYEYAERYKAVADKYDVDLSEID HQLMILVNYNEDSNKAKQETRAFISDYVLEMHPNENFENKLEEIIAENAVGNYTECITA AKLAIEKCGAKSVLLSFEPMNDLMSQKNVINIVDDNIKKYHTEYT" CDS complement(4441..4464) /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" terminator 5296..5323 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" terminator 5415..5501 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" primer_bind 5654..5671 /label=pBAD Reverse /note="For vectors with E. coli araBAD promoter, reverse primer" CDS complement(5717..6577) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 6340..6359 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6578..6682) /gene="bla" /label=AmpR promoter primer_bind 6750..6768 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" ORIGIN 1 tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg gttcgttgtt tttgcgtgag 61 ccatgagaac gaaccattga gatcatgctt actttgcatg tcactcaaaa attttgcctc 121 aaaactggtg agctgaattt ttgcagttaa agcatcgtgt agtgtttttc ttagtccgtt 181 acgtaggtag gaatctgatg taatggttgt tggtattttg tcaccattca tttttatctg 241 gttgttctca agttcggtta cgagatccat ttgtctatct agttcaactt ggaaaatcaa 301 cgtatcagtc gggcggcctc gcttatcaac caccaatttc atattgctgt aagtgtttaa 361 atctttactt attggtttca aaacccattg gttaagcctt ttaaactcat ggtagttatt 421 ttcaagcatt aacatgaact taaattcatc aaggctaatc tctatatttg ccttgtgagt 481 tttcttttgt gttagttctt ttaataacca ctcataaatc ctcatagagt atttgttttc 541 aaaagactta acatgttcca gattatattt tatgaatttt tttaactgga aaagataagg 601 caatatctct tcactaaaaa ctaattctaa tttttcgctt gagaacttgg catagtttgt 661 ccactggaaa atctcaaagc ctttaaccaa aggattcctg atttccacag ttctcgtcat 721 cagctctctg gttgctttag ctaatacacc ataagcattt tccctactga tgttcatcat 781 ctgagcgtat tggttataag tgaacgatac cgtccgttct ttccttgtag ggttttcaat 841 cgtggggttg agtagtgcca cacagcataa aattagcttg gtttcatgct ccgttaagtc 901 atagcgacta atcgctagtt catttgcttt gaaaacaact aattcagaca tacatctcaa 961 ttggtctagg taattttaat cactatacca attgagatgg gctagtcaat gataattact 1021 agtccttttc ctttgagttg tgggtatctg taaattctgc tagacctttg ctggaaaact 1081 tgtaaattct gctagaccct ctgtaaattc cgctagacct ttgtgtgttt tttttgttta 1141 tattcaagtg gttataattt atagaataaa gaaagaataa aaaaagataa aaagaataga 1201 tcccagccct gtgtataact cactacttta gtcagttccg cagtattaca aaaggatgtc 1261 gcaaacgctg tttgctcctc tacaaaacag accttaaaac cctaaaggct taagtagcac 1321 cctcgcaagc tcgggcaaat cgctgaatat tccttttgtc tccgaccatc aggcacctga 1381 gtcgctgtct ttttcgtgac attcagttcg ctgcgctcac ggctctggca gtgaatgggg 1441 gtaaatggca ctacaggcgc cttttatgga ttcatgcaag gaaactaccc ataatacaag 1501 aaaagcccgt cacgggcttc tcagggcgtt ttatggcggg tctgctatgt ggtgctatct 1561 gactttttgc tgttcagcag ttcctgccct ctgattttcc agtctgacca cttcggatta 1621 tcccgtgaca ggtcattcag actggctaat gcacccagta aggcagcggt atcatcaaca 1681 ggcttacccg tcgtacggtc aagaggacat ccgggcaaat gcaacgaaag gctcagtcga 1741 aagactgggc ctttcgttgc gctgcggaga cttagggacc ctaaaacaag atacattgta 1801 tgaaaataca agaaagtttg ttcattaggc accccgggct ttactcgtaa agcttccggc 1861 gcgtatgttg tgtcgaccga cctgcaggtg cagtaaggag gaaaaaaaaa tgaaatttgg 1921 aaactttttg cttacatacc aacctcccca attttcccaa acagaggtaa tgaaacgttt 1981 ggttaaatta ggtcgcatct ctgaggagtg tggttttgat accgtatggt tactggagca 2041 tcatttcacg gagtttggtt tgcttggtaa cccttatgtc gctgctgcat atttacttgg 2101 cgcgactaaa aaattgaatg taggaactgc cgctattgtt cttcccacag cccatccagt 2161 acgccaactt gaagatgtga atttattgga tcaaatgtca aaaggacgat ttcggtttgg 2221 tatttgccga gggctttaca acaaggactt tcgcgtattc ggcacagata tgaataacag 2281 tcgcgcctta gcggaatgct ggtacgggct gataaagaat ggcatgacag agggatatat 2341 ggaagctgat aatgaacata tcaagttcca taaggtaaaa gtaaaccccg cggcgtatag 2401 cagaggtggc gcaccggttt atgtggtggc tgaatcagct tcgacgactg agtgggctgc 2461 tcaatttggc ctaccgatga tattaagttg gattataaat actaacgaaa agaaagcaca 2521 acttgagctt tataatgaag tggctcaaga atatgggcac gatattcata atatcgacca 2581 ttgcttatca tatataacat ctgtagatca tgactcaatt aaagcgaaag agatttgccg 2641 gaaatttctg gggcattggt atgattctta tgtgaatgct acgactattt ttgatgattc 2701 agaccaaaca agaggttatg atttcaataa agggcagtgg cgtgactttg tattaaaagg 2761 acataaagat actaatcgcc gtattgatta cagttacgaa atcaatcccg tgggaacgcc 2821 gcaggaatgt attgacataa ttcaaaaaga cattgatgct acaggaatat caaatatttg 2881 ttgtggattt gaagctaatg gaacagtaga cgaaattatt gcttccatga agctcttcca 2941 gtctgatgtc atgccatttc ttaaagaaaa acaacgttcg ctattatatt atggcggtgg 3001 cggtagcggc ggtggcggta gcggcggtgg cggtagcggc ggtggcggta gcaaatttgg 3061 attgttcttc cttaacttca tcaattcaac aactgttcaa gaacagagta tagttcgcat 3121 gcaggaaata acggagtatg ttgataagtt gaattttgaa cagattttag tgtatgaaaa 3181 tcatttttca gataatggtg ttgtcggcgc tcctctgact gtttctggtt ttctgctcgg 3241 tttaacagag aaaattaaaa ttggttcatt aaatcacatc attacaactc atcatcctgt 3301 ccgcatagcg gaggaagctt gcttattgga tcagttaagt gaagggagat ttattttagg 3361 gtttagtgat tgcgaaaaaa aagatgaaat gcattttttt aatcgcccgg ttgaatatca 3421 acagcaacta tttgaagagt gttatgaaat cattaacgat gctttaacaa caggctattg 3481 taatccagat aacgattttt atagcttccc taaaatatct gtaaatcccc atgcttatac 3541 gccaggcgga cctcggaaat atgtaacagc aaccagtcat catattgttg agtgggcggc 3601 caaaaaaggt attcctctca tctttaagtg ggatgattct aatgatgtta gatatgaata 3661 tgctgaaaga tataaagccg ttgcggataa atatgacgtt gacctatcag agatagacca 3721 tcagttaatg atattagtta actataacga agatagtaat aaagctaaac aagagacgcg 3781 tgcatttatt agtgattatg ttcttgaaat gcaccctaat gaaaatttcg aaaataaact 3841 tgaagaaata attgcagaaa acgctgtcgg aaattatacg gagtgtataa ctgcggctaa 3901 gttggcaatt gaaaagtgtg gtgcgaaaag tgtattgctg tcctttgaac caatgaatga 3961 tttgatgagc caaaaaaatg taatcaatat tgttgatgat aatattaaga agtaccacac 4021 ggaatatacc taaacggagc caatgtacgc aaaaaacccc gcttcagcgg ggttttttcg 4081 ccaaaaaaaa ccccgccctg tcaggggcgg ggtttttttt tagggcccta ctgactgtta 4141 gcgtttcggc gccggataat gcagcgtggt gatcagacca tccgcaaccg tgctatggtg 4201 atgcaccagt tctgccaggg tattgaagcg agattcacta gaaacgtaca gtttgccatc 4261 gctggcggtg ttgatacggt agtgatacac gcgaccttca taacgcagag aaatgctgcg 4321 ctggcccggg ctactttcac tttcacgaac cagaaaagaa ccattaatgc cagagctcag 4381 caggtattcg gccgcgttac ggctcacagg gccatgatac cagctgtgtt tttccagact 4441 cttgtcatcg tcgtccttgt agtctacgaa ttttaccctc gcttccacga caacaccgat 4501 aatcttgcag tttccgttga taggagtcat aggccatgaa ggattcaggc ctttcaggta 4561 cttctgaccg ccatctatga ccagtttctt gaatgttgct tcgttcgcgt cagtcagttt 4621 ggctacaaca aggcttccat tcactggctc ccgtccagta tctactaaca ccatatgacc 4681 ttcagggatg ctttgaccta caggtgaggt catggaatca ccttcaacct tcagccagaa 4741 tccattgcct aataagttaa cgtcactgtc ataccattca tcaatgtcct tgatatcgta 4801 gggttcacaa gcttcacacc acgaacgagc tctaaccatg ctaatcaatg gatatttccc 4861 tttgggctca acgtgcccaa caaatctaac attcgaatca gaggtgccat tgagcagcca 4921 gtcaacactt acgccaagag ctgacgcaag ttctggtaaa aagcgtggtc gcttagtttt 4981 accgttttcg agctgctcta tagactgctg ggtagtcccc accttttgag caagttcagc 5041 ctggttaagt ccaagctgaa ttcttttgct ttttaccctg ctgctgatac tcattttttt 5101 ttcctcctta ctctagtaaa agttaaacaa aattatttgt agagggaaac cgttgtggtc 5161 tccctgaata tagataccga gccttatgca tgcccgtaaa gttatccagc aaccactcat 5221 agacctaggg cagcagatag ggacgacgtg gtgttagctg tgctctagaa tcctcagcga 5281 gtttgtagaa acgcaaaaag gccatccgtc aggatggcct tctgcttaat ttgatgcctg 5341 gcagtttatg gcgggcgtcc tgcccgccac cctccgggcc gttgcttcgc aacgttcaaa 5401 tccgctcccg gcggatttgt cctactcagg agagcgttca ccgacaaaca acagataaaa 5461 cgaaaggccc agtctttcga ctgagccttt cgttttattt gatgcctggc agttccctac 5521 tctcgcatgg ggagacccca cactaccatc ggcgctacgg cgtttcactt ctgagttcgg 5581 catggggtca ggtgggacca ccgcgctact gccgccaggc aaattctgtt ttatcagacc 5641 gcttctgcgt tctgatttaa tctgtatcag gctgaaaatc ttctctcatc cgccaaaaca 5701 gccatgccca tggcgtttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc 5761 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg 5821 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag 5881 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt 5941 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag 6001 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt 6061 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca 6121 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg 6181 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat 6241 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta 6301 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca 6361 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct 6421 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat 6481 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa 6541 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt 6601 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa 6661 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa 6721 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc ttaggccaaa 6781 taggccgt //