LOCUS Exported 4075 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS TLD_026 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4075) AUTHORS Deans TL, Cantor CR, Collins JJ TITLE A tunable genetic switch based on RNAi and repressor proteins for regulating gene expression in mammalian cells. JOURNAL Cell. 2007 Jul 27. 130(2):363-72. PUBMED 17662949 REFERENCE 2 (bases 1 to 4075) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4075 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(29..51) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 151..170 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 364..386 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 378..395 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 379..395 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" enhancer 462..765 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 766..969 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 920..940 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" CDS 1024..1641 /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 DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESG" primer_bind complement(1097..1116) /label=Tet-R /note="Tetracycline resistance gene, reverse primer" polyA_signal 1686..1807 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1723..1742) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1777..1796 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(1854..1870) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1854..1870) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1867..1889) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1878..1894 /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)." promoter complement(1902..1932) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1947..1968 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(2085..2102) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(2256..2844) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2336..2355) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(3015..3875) /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 3638..3657 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(3876..3980) /gene="bla" /label=AmpR promoter primer_bind 4048..4066 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" ORIGIN 1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca 61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg 121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc 181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc 241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat 301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt 361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt ctagttatta atagtaatca 421 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta 481 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat 541 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg 601 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac 661 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt 721 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg 781 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc 841 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt 901 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata 961 agcagagctg gtttagtgaa ccgtcagatc ggatcctcta gagtcgacct gcaggccgcc 1021 accatgtcta gattagataa aagtaaagtg attaacagcg cattagagct gcttaatgag 1081 gtcggaatcg aaggtttaac aacccgtaaa ctcgcccaga agctaggtgt agagcagcct 1141 acattgtatt ggcatgtaaa aaataagcgg gctttgctcg acgccttagc cattgagatg 1201 ttagataggc accatactca cttttgccct ttagaagggg aaagctggca agatttttta 1261 cgtaataacg ctaaaagttt tagatgtgct ttactaagtc atcgcgatgg agcaaaagta 1321 catttaggta cacggcctac agaaaaacag tatgaaactc tcgaaaatca attagccttt 1381 ttatgccaac aaggtttttc actagagaat gcattatatg cactcagcgc tgtggggcat 1441 tttactttag gttgcgtatt ggaagatcaa gagcatcaag tcgctaaaga agaaagggaa 1501 acacctacta ctgatagtat gccgccatta ttacgacaag ctatcgaatt atttgatcac 1561 caaggtgcag agccagcctt cttattcggc cttgaattga tcatatgcgg attagaaaaa 1621 caacttaaat gtgaaagtgg gtccgcgtac agcggatccc gggaattcag atcttattaa 1681 agcagaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt 1741 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg 1801 tatcttatca tgtctggtcg aatccatcac actggaagct tggcgtaatc atggtcatag 1861 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc 1921 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc 1981 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa 2041 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg 2101 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg 2161 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag 2221 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac 2281 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga 2341 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt 2401 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc 2461 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc 2521 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta 2581 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat 2641 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca 2701 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct 2761 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt 2821 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct 2881 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc 2941 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa 3001 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta 3061 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc 3121 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat 3181 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta 3241 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt 3301 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt 3361 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg 3421 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc 3481 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc 3541 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg 3601 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga 3661 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta 3721 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct 3781 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag 3841 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga 3901 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat 3961 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc 4021 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtc //