LOCUS Exported 2932 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pTREtight2 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 2932) TITLE pTREtight with high copy E. coli origin REFERENCE 2 (bases 1 to 2932) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..2932 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(1..25) /label=LNCX /note="Human CMV promoter, forward primer" protein_bind 72..90 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 108..126 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 143..161 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 179..197 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 215..233 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 250..268 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 286..304 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" promoter complement(321..351) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 366..387 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(504..521) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(675..1263) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(755..774) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(1434..2294) /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 2057..2076 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(2295..2399) /gene="bla" /label=AmpR promoter primer_bind 2467..2485 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(2523..2545) /label=pGEX 3' /note="pGEX vectors, reverse primer" polyA_signal 2648..2729 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind 2673..2692 /label=SV40pA-R /note="SV40 polyA, reverse primer" promoter complement(2930..312) /label=tight TRE promoter /note="Tet-responsive promoter PTight, consisting of seven tet operator sequences followed by the minimal CMV promoter" ORIGIN 1 gatctgacgg ttcactaaac gagctctgct tatataggcc tcccaccgta cacgcctacc 61 tcgacatacg ttctctatca ctgataggga gtaaactcga catacgttct ctatcactga 121 tagggataaa ctcgacatac gttctctatc actgataggg agtaaactcg acatacgttc 181 tctatcactg atagggagta aactcgacat acgttctcta tcactgatag ggagtaaact 241 cgacatcgtt ctctatcact gatagggagt aaactcgaca tacgttctct atcactgata 301 gggagtaaac tcgagccaca caacatacga gccggaagca taaagtgtaa agcctggggt 361 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg 421 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg 481 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg 541 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat 601 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc 661 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc 721 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga 781 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt 841 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg 901 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc 961 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg 1021 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc 1081 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg 1141 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc 1201 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct 1261 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt 1321 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa 1381 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa 1441 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc 1501 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct 1561 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca 1621 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt 1681 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt 1741 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc 1801 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc 1861 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt 1921 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact 1981 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc 2041 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt 2101 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg 2161 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct 2221 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa 2281 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt 2341 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc 2401 acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc 2461 tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa 2521 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg 2581 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac 2641 tctcgaggca gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg 2701 taaccattat aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc 2761 aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta 2821 tggctgatta tgatcctcta gagatatcgt cgacaagctt atcgatgcgg ccgcgctagc 2881 acgcgtcagc tgactagagg atccccgggt accgagctcg aattctccag gc //