LOCUS Exported 6697 bp ds-DNA circular SYN 30-OCT-2018 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6697) AUTHORS MReijns TITLE Direct Submission JOURNAL Exported Oct 30, 2018 from SnapGene Viewer 4.2.4 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6697 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 183..211 /label=tac promoter /note="strong E. coli promoter; hybrid between the trp and lac UV5 promoters" protein_bind 219..235 /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 258..911 /codon_start=1 /product="glutathione S-transferase from Schistosoma japonicum" /label=GST /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK" CDS 918..941 /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /label=HRV 3C site /translation="LEVLFQGP" protein_bind 966..1090 /gene="mutant version of attR" /label=attR1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 1115..1145 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" CDS 1199..1858 /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" CDS 2200..2505 /codon_start=1 /gene="ccdB" /product="CcdB, a bacterial toxin that poisons DNA gyrase" /label=ccdB /note="Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains." /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI" protein_bind complement(2546..2670) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 3000..3104 /gene="bla" /label=AmpR promoter CDS 3105..3965 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" rep_origin 4136..4724 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" promoter 4968..5045 /gene="lacI (mutant)" /label=lacIq promoter /note="In the lacIq allele, a single base change in the promoter boosts expression of the lacI gene about 10-fold." CDS 5046..6128 /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 YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR ALADSLMQLARQVSRLESGQ" protein_bind 6141..6162 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 6177..6207 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6215..6231 /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)." primer_bind 6239..6255 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" CDS 6251..6697 /codon_start=1 /gene="lacZ fragment" /product="LacZ-alpha fragment of beta-galactosidase" /label=lacZ-alpha /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGI" primer_bind complement(6271..6287) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" ORIGIN 1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg 61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt 121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc 181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca 241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc 301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc 361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc 421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata 481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc 541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact 601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag 661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt 721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa 781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat 841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc 901 atcctccaaa atcggatctg gaagttctgt tccaggggcc cctgggatcc ccggaattcc 961 catcaacaag tttgtacaaa aaagctgaac gagaaacgta aaatgatata aatatcaata 1021 tattaaatta gattttgcat aaaaaacaga ctacataata ctgtaaaaca caacatatcc 1081 agtcactatg gcggccgcat taggcacccc aggctttaca ctttatgctt ccggctcgta 1141 taatgtgtgg attttgagtt aggatccgtc gagattttca ggagctaagg aagctaaaat 1201 ggagaaaaaa atcactggat ataccaccgt tgatatatcc caatggcatc gtaaagaaca 1261 ttttgaggca tttcagtcag ttgctcaatg tacctataac cagaccgttc agctggatat 1321 tacggccttt ttaaagaccg taaagaaaaa taagcacaag ttttatccgg cctttattca 1381 cattcttgcc cgcctgatga atgctcatcc ggaattccgt atggcaatga aagacggtga 1441 gctggtgata tgggatagtg ttcacccttg ttacaccgtt ttccatgagc aaactgaaac 1501 gttttcatcg ctctggagtg aataccacga cgatttccgg cagtttctac acatatattc 1561 gcaagatgtg gcgtgttacg gtgaaaacct ggcctatttc cctaaagggt ttattgagaa 1621 tatgtttttc gtctcagcca atccctgggt gagtttcacc agttttgatt taaacgtggc 1681 caatatggac aacttcttcg cccccgtttt caccatgggc aaatattata cgcaaggcga 1741 caaggtgctg atgccgctgg cgattcaggt tcatcatgcc gtttgtgatg gcttccatgt 1801 cggcagaatg cttaatgaat tacaacagta ctgcgatgag tggcagggcg gggcgtaaag 1861 atctggatcc ggcttactaa aagccagata acagtatgcg tatttgcgcg ctgatttttg 1921 cggtataaga atatatactg atatgtatac ccgaagtatg tcaaaaagag gtatgctatg 1981 aagcagcgta ttacagtgac agttgacagc gacagctatc agttgctcaa ggcatatatg 2041 atgtcaatat ctccggtctg gtaagcacaa ccatgcagaa tgaagcccgt cgtctgcgtg 2101 ccgaacgctg gaaagcggaa aatcaggaag ggatggctga ggtcgcccgg tttattgaaa 2161 tgaacggctc ttttgctgac gagaacaggg gctggtgaaa tgcagtttaa ggtttacacc 2221 tataaaagag agagccgtta tcgtctgttt gtggatgtac agagtgatat tattgacacg 2281 cccgggcgac ggatggtgat ccccctggcc agtgcacgtc tgctgtcaga taaagtctcc 2341 cgtgaacttt acccggtggt gcatatcggg gatgaaagct ggcgcatgat gaccaccgat 2401 atggccagtg tgccggtctc cgttatcggg gaagaagtgg ctgatctcag ccaccgcgaa 2461 aatgacatca aaaacgccat taacctgatg ttctggggaa tataaatgtc aggctccctt 2521 atacacagcc agtctgcagg tcgaccatag tgactggata tgttgtgttt tacagtatta 2581 tgtagtctgt tttttatgca aaatctaatt taatatattg atatttatat cattttacgt 2641 ttctcgttca gctttcttgt acaaagtggt tgatgggtcg actcgagcgg ccgcatcgtg 2701 actgactgac gatctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc 2761 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc 2821 agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg 2881 atagcggagt gtataattct tgaagacgaa agggcctcgt gatacgccta tttttatagg 2941 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc 3001 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac 3061 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt 3121 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag 3181 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg 3241 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa 3301 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc 3361 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag 3421 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa 3481 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc 3541 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg 3601 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgca gcaatggcaa 3661 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa 3721 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg 3781 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag 3841 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg 3901 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt 3961 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt 4021 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac 4081 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag 4141 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg 4201 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca 4261 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga 4321 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca 4381 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc 4441 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca 4501 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa 4561 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc 4621 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc 4681 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg 4741 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat 4801 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca 4861 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt 4921 attttctcct tacgcatctg tgcggtattt cacaccgcat aaattccgac accatcgaat 4981 ggtgcaaaac ctttcgcggt atggcatgat agcgcccgga agagagtcaa ttcagggtgg 5041 tgaatgtgaa accagtaacg ttatacgatg tcgcagagta tgccggtgtc tcttatcaga 5101 ccgtttcccg cgtggtgaac caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg 5161 aagcggcgat ggcggagctg aattacattc ccaaccgcgt ggcacaacaa ctggcgggca 5221 aacagtcgtt gctgattggc gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa 5281 ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg 5341 tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg 5401 tcagtgggct gatcattaac tatccgctgg atgaccagga tgccattgct gtggaagctg 5461 cctgcactaa tgttccggcg ttatttcttg atgtctctga ccagacaccc atcaacagta 5521 ttattttctc ccatgaagac ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc 5581 accagcaaat cgcgctgtta gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg 5641 ctggctggca taaatatctc actcgcaatc aaattcagcc gatagcggaa cgggaaggcg 5701 actggagtgc catgtccggt tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc 5761 ccactgcgat gctggttgcc aacgatcaga tggcgctggg cgcaatgcgc gccattaccg 5821 agtccgggct gcgcgttggt gcggatatct cggtagtggg atacgacgat accgaagaca 5881 gctcatgtta tatcccgccg tcaaccacca tcaaacagga ttttcgcctg ctggggcaaa 5941 ccagcgtgga ccgcttgctg caactctctc agggccaggc ggtgaagggc aatcagctgt 6001 tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc caatacgcaa accgcctctc 6061 cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg 6121 ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta 6181 cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca 6241 ggaaacagct atgaccatga ttacggattc actggccgtc gttttacaac gtcgtgactg 6301 ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg 6361 gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg 6421 cgaatggcgc tttgcctggt ttccggcacc agaagcggtg ccggaaagct ggctggagtg 6481 cgatcttcct gaggccgata ctgtcgtcgt cccctcaaac tggcagatgc acggttacga 6541 tgcgcccatc tacaccaacg taacctatcc cattacggtc aatccgccgt ttgttcccac 6601 ggagaatccg acgggttgtt actcgctcac atttaatgtt gatgaaagct ggctacagga 6661 aggccagacg cgaattattt ttgatggcgt tggaatt //