LOCUS Exported 6664 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS repressilator (pZS1-lTlrLLtCL) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6664) AUTHORS Elowitz MB, Leibler S TITLE A synthetic oscillatory network of transcriptional regulators. JOURNAL Nature. 2000 Jan 20. 403(6767):335-8. PUBMED 10659856 REFERENCE 2 (bases 1 to 6664) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6664 /organism="synthetic DNA construct" /mol_type="other DNA" protein_bind 8..24 /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)." protein_bind 31..47 /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)." RBS 89..100 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" CDS 107..727 /codon_start=1 /gene="tetR from transposon Tn10" /product="tetracycline repressor TetR" /label=TetR /note="TetR binds to the tetracycline operator tetO to inhibit transcription. This inhibition can be relieved by adding tetracycline or doxycycline." /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS" primer_bind complement(180..199) /label=Tet-R /note="Tetracycline resistance gene, reverse primer" terminator 775..861 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" RBS 968..979 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" CDS 997..2076 /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 complement(1016..1035) /label=LacI-R /note="LacI, reverse primer" terminator 2141..2227 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" promoter 2258..2331 /label=PLtetO-1 promoter /note="modified phage lambda PL promoter with tet operator sites (Lutz and Bujard, 1997)" protein_bind 2258..2276 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 2283..2301 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" RBS 2341..2352 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" CDS 2359..3069 /codon_start=1 /gene="cI" /product="phage lambda repressor" /label=-lambda repressor /translation="MSTKKKPLTQEQLEDARRLKAIYEKKKNELGLSQESVADKMGMGQ SGVGALFNGINALNAYNAALLAKILKVSVEEFSPSIAREIYEMYEAVSMQPSLRSEYEY PVFSHVQAGMFSPELRTFTKGDAERWVSTTKKASDSAFWLEVEGNSMTAPTGSKPSFPD GMLILVDPEQAVEPGDFCIARLGGDEFTFKKLIRDSGQVFLQPLNPQYPMIPCNESCSV VGKVIASQWPEETFG" terminator 3141..3227 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" CDS complement(3762..4712) /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 4760..4982 /label=pSC101 ori /note="low-copy replication origin that requires the Rep101 protein" terminator 5474..5568 /label=lambda t0 terminator /note="transcription terminator from phage lambda" CDS complement(5598..6458) /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 6221..6240 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6459..6563) /gene="bla" /label=AmpR promoter primer_bind 6631..6649 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" ORIGIN 1 tcgagaattg tgagcggata acaattgaca ttgtgagcgg ataacaagat actgagcaca 61 tcagcaggac gcactgaccg aattcattaa agaggagaaa ggtaccatgt ccagattaga 121 taaaagtaaa gtgattaaca gcgcattaga gctgcttaat gaggtcggaa tcgaaggttt 181 aacaacccgt aaactcgccc agaagctagg tgtagagcag cctacattgt attggcatgt 241 aaaaaataag cgggctttgc tcgacgcctt agccattgag atgttagata ggcaccatac 301 tcacttttgc cctttagaag gggaaagctg gcaagatttt ttacgtaata acgctaaaag 361 ttttagatgt gctttactaa gtcatcgcga tggagcaaaa gtacatttag gtacacggcc 421 tacagaaaaa cagtatgaaa ctctcgaaaa tcaattagcc tttttatgcc aacaaggttt 481 ttcactagag aatgcattat atgcactcag cgctgtgggg cattttactt taggttgcgt 541 attggaagat caagagcatc aagtcgctaa agaagaaagg gaaacaccta ctactgatag 601 tatgccgcca ttattacgac aagctatcga attatttgat caccaaggtg cagagccagc 661 cttcttattc ggccttgaat tgatcatatg cggattagaa aaacaactta aatgtgaaag 721 tgggtctgca gcaaacgacg aaaactacgc tttagcagct taatctagag gcatcaaata 781 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac 841 gctctcctga gtaggacaaa tccgccgccc tagacctagc tgcaggtcga ggataaatat 901 ctaacaccgt gcgtgttgac tattttacct ctggcggtga taatggttgc atgtactaga 961 attcattaaa gaggagaaag gtaccatatg gtgaatgtga aaccagtaac gttatacgat 1021 gtcgcagagt atgccggtgt ctcttatcag accgtttccc gcgtggtgaa ccaggccagc 1081 cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct gaattacatt 1141 cccaaccgcg tggcacaaca actggcgggc aaacagtcgt tgctgattgg cgttgccacc 1201 tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat 1261 caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa 1321 gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg 1381 gatgaccagg atgccattgc tgtggaagct gcctgcacta atgttccggc gttatttctt 1441 gatgtctctg accagacacc catcaacagt attattttct cccatgaaga cggtacgcga 1501 ctgggcgtgg agcatctggt cgcattgggt caccagcaaa tcgcgctgtt agcgggccca 1561 ttaagttctg tctcggcgcg tctgcgtctg gctggctggc ataaatatct cactcgcaat 1621 caaattcagc cgatagcgga acgggaaggc gactggagtg ccatgtccgg ttttcaacaa 1681 accatgcaaa tgctgaatga gggcatcgtt cccactgcga tgctggttgc caacgatcag 1741 atggcgctgg gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg tgcggatatc 1801 tcggtagtgg gatacgacga taccgaagac agctcatgtt atatcccgcc gttaaccacc 1861 atcaaacagg attttcgcct gctggggcaa accagcgtgg accgcttgct gcaactctct 1921 cagggccagg cggtgaaggg caatcagctg ttgcccgtct cactggtgaa aagaaaaacc 1981 accctggcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag 2041 ctggcacgac aggtttcccg actggaaagc gggcaggcag caaacgacga aaactacgct 2101 ttagcagctt aaaagcttaa ttagctgagt ctagaggcat caaataaaac gaaaggctca 2161 gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag 2221 gacaaatccg ccgccctaga cctagctgca ggtcgagtcc ctatcagtga tagagattga 2281 catccctatc agtgatagag atactgagca catcagcagg acgcactgac cgaattcatt 2341 aaagaggaga aaggtaccat gagcacaaaa aagaaaccat taacacaaga gcagcttgag 2401 gacgcacgtc gccttaaagc aatttatgaa aaaaagaaaa atgaacttgg cttatcccag 2461 gaatctgtcg cagacaagat ggggatgggg cagtcaggcg ttggtgcttt atttaatggc 2521 atcaatgcat taaatgctta taacgccgca ttgcttgcaa aaattctcaa agttagcgtt 2581 gaagaattta gcccttcaat cgccagagaa atctacgaga tgtatgaagc ggttagtatg 2641 cagccgtcac ttagaagtga gtatgagtac cctgtttttt ctcatgttca ggcagggatg 2701 ttctcacctg agcttagaac ctttaccaaa ggtgatgcgg agagatgggt aagcacaacc 2761 aaaaaagcca gtgattctgc attctggctt gaggttgaag gtaattccat gaccgcacca 2821 acaggctcca agccaagctt tcctgacgga atgttaattc tcgttgaccc tgagcaggct 2881 gttgagccag gtgatttctg catagccaga cttgggggtg atgagtttac cttcaagaaa 2941 ctgatcaggg atagcggtca ggtgttttta caaccactaa acccacagta cccaatgatc 3001 ccatgcaatg agagttgttc cgttgtgggg aaagttatcg ctagtcagtg gcctgaagag 3061 acgtttggcg cagcaaacga cgaaaactac gctttagcag cttgatctag aggatcccat 3121 ggtacgcgtg ctagaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc 3181 gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg ccgccctaga 3241 cctagggtac gggttttgct gcccgcaaac gggctgttct ggtgttgcta gtttgttatc 3301 agaatcgcag atccggcttc aggtttgccg gctgaaagcg ctatttcttc cagaattgcc 3361 atgatttttt ccccacggga ggcgtcactg gctcccgtgt tgtcggcagc tttgattcga 3421 taagcagcat cgcctgtttc aggctgtcta tgtgtgactg ttgagctgta acaagttgtc 3481 tcaggtgttc aatttcatgt tctagttgct ttgttttact ggtttcacct gttctattag 3541 gtgttacatg ctgttcatct gttacattgt cgatctgttc atggtgaaca gctttaaatg 3601 caccaaaaac tcgtaaaagc tctgatgtat ctatcttttt tacaccgttt tcatctgtgc 3661 atatggacag ttttcccttt gatatctaac ggtgaacagt tgttctactt ttgtttgtta 3721 gtcttgatgc ttcactgata gatacaagag ccataagaac ctcagatcct tccgtattta 3781 gccagtatgt tctctagtgt ggttcgttgt ttttgcgtga gccatgagaa cgaaccattg 3841 agatcatgct tactttgcat gtcactcaaa aattttgcct caaaactggt gagctgaatt 3901 tttgcagtta aagcatcgtg tagtgttttt cttagtccgt tacgtaggta ggaatctgat 3961 gtaatggttg ttggtatttt gtcaccattc atttttatct ggttgttctc aagttcggtt 4021 acgagatcca tttgtctatc tagttcaact tggaaaatca acgtatcagt cgggcggcct 4081 cgcttatcaa ccaccaattt catattgctg taagtgttta aatctttact tattggtttc 4141 aaaacccatt ggttaagcct tttaaactca tggtagttat tttcaagcat taacatgaac 4201 ttaaattcat caaggctaat ctctatattt gccttgtgag ttttcttttg tgttagttct 4261 tttaataacc actcataaat cctcatagag tatttgtttt caaaagactt aacatgttcc 4321 agattatatt ttatgaattt ttttaactgg aaaagataag gcaatatctc ttcactaaaa 4381 actaattcta atttttcgct tgagaacttg gcatagtttg tccactggaa aatctcaaag 4441 cctttaacca aaggattcct gatttccaca gttctcgtca tcagctctct ggttgcttta 4501 gctaatacac cataagcatt ttccctactg atgttcatca tctgagcgta ttggttataa 4561 gtgaacgata ccgtccgttc tttccttgta gggttttcaa tcgtggggtt gagtagtgcc 4621 acacagcata aaattagctt ggtttcatgc tccgttaagt catagcgact aatcgctagt 4681 tcatttgctt tgaaaacaac taattcagac atacatctca attggtctag gtgattttaa 4741 tcactatacc aattgagatg ggctagtcaa tgataattac tagtcctttt cctttgagtt 4801 gtgggtatct gtaaattctg ctagaccttt gctggaaaac ttgtaaattc tgctagaccc 4861 tctgtaaatt ccgctagacc tttgtgtgtt ttttttgttt atattcaagt ggttataatt 4921 tatagaataa agaaagaata aaaaaagata aaaagaatag atcccagccc tgtgtataac 4981 tcactacttt agtcagttcc gcagtattac aaaaggatgt cgcaaacgct gtttgctcct 5041 ctacaaaaca gaccttaaaa ccctaaaggc ttaagtagca ccctcgcaag ctcgggcaaa 5101 tcgctgaata ttccttttgt ctccgaccat caggcacctg agtcgctgtc tttttcgtga 5161 cattcagttc gctgcgctca cggctctggc agtgaatggg ggtaaatggc actacaggcg 5221 ccttttatgg attcatgcaa ggaaactacc cataatacaa gaaaagcccg tcacgggctt 5281 ctcagggcgt tttatggcgg gtctgctatg tggtgctatc tgactttttg ctgttcagca 5341 gttcctgccc tctgattttc cagtctgacc acttcggatt atcccgtgac aggtcattca 5401 gactggctaa tgcacccagt aaggcagcgg tatcatcaac aggcttaccc gtcttactgt 5461 ccctagtgct tggattctca ccaataaaaa acgcccggcg gcaaccgagc gttctgaaca 5521 aatccagatg gagttctgag gtcattactg gatctatcaa caggagtcca agcgagctcg 5581 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt 5641 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag 5701 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca 5761 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact 5821 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca 5881 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg 5941 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc 6001 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg 6061 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca 6121 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt 6181 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc 6241 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc 6301 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca 6361 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa 6421 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat 6481 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa 6541 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa 6601 accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctt 6661 cacc //