LOCUS Exported 6475 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION ""Burdensome"" GFP expressing plasmid carrying axe/txe toxin-antitoxin for plasmid stabilisation. ACCESSION . VERSION . KEYWORDS pUC-GFP-AT SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6475) AUTHORS Fedorec AJH, Ozdemir T, Doshi A, Ho YK, Rosa L, Rutter J, Velazquez O, Pinheiro VB, Danino T, Barnes CP TITLE Two New Plasmid Post-segregational Killing Mechanisms for the Implementation of Synthetic Gene Networks in Escherichia coli. JOURNAL iScience. 2019 Apr 26;14:323-334. doi: 10.1016/j.isci.2019.03.019. Epub 2019 Mar 22. PUBMED 30954530 REFERENCE 2 (bases 1 to 6475) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6475 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 18..89 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 94..140 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 158..206 /note="synthetic polyadenylation signal" polyA_signal 2213..2334 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2224..2243) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2278..2297 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 2449..2495 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 2525..2661 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 2706..2777 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(3003..3591) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3083..3102) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 4211..4279 /label=recA (Delta-LexA) promoter /note="constitutively active E. coli recA promoter lacking the LexA binding site (Brent and Ptashne, 1981)" CDS 4296..5006 /codon_start=1 /product="green fluorescent protein with peak excitation at 510 nm and peak emission at 521 nm" /label=daGFP /translation="MTALTEGAKLFEKEIPYITELEGDVEGMKFIIKGEGTGDATTGTI KAKYICTTGDLPVPWATLVSTLSYGVQCFAKYPSHIKDFFKSAMPEGYTQERTISFEGD GVYKTRAMVTYERGSIYNRVTLTGENFKKDGHILRKNVAFQCPPSILYILPDTVNNGIR VEFNQAYDIEGVTEKLVTKCSQMNRPLAGSAAVHIPRYHHITYHTKLSKDRDERRDHMC LVEVVKAVDLDTYQ" terminator 5013..5148 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" CDS 5405..6199 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(5459..5478) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 6069..6088 /label=Neo-F /note="Neomycin resistance gene, forward primer" terminator 6212..6348 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" ORIGIN 1 gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct tccctctagt 61 ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga cccgtttaga 121 ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc aacacacatc 181 catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc cagatctttg 241 tcgatcctac catccactcg acacacccgc cagcggccgc tgccaagctt ccgagctcga 301 tacttctttc aaattcaagt taaatttata ttccattact tctcctccaa tatataaaaa 361 ttacttaact gtatccaaat ttaatatgta aataaatcat taatttttag ctaaattcta 421 tgagaagttt atttttgttg tttttgctaa ttgactaaaa ctagtacaaa aaatttaaaa 481 tagattatta aacctattgc ttattctatt cagtcatata tataagcgtc atcctttagt 541 aactaacatc ccacactaca atagtattat aacatatcta ttcaacttca aattgttgaa 601 atttaagcca tttatattta cttttcctca actttagaaa tgattttact aaccttatat 661 ccgtaatacg cgtaacaaac acattcatta ataacaatca acactaagct atattagctt 721 tttaaatgac tagggtaaat aaaataagga tcaaaaactt ttaagtttct gaccctttcc 781 ttacttccga ttggttaata gtgatctttt gcagaataaa taaatatcgt ttcattttca 841 actctatata tcagtctatg ttcatctgta attcttctgg accattttcc agataaatca 901 tgctttaatg gctcaggttt tcctaatcca gcaaaggggg aacgatcgat atcttttatt 961 aacttgttaa tcttttttat attgcttttg tttccttgct catgccaata aagataatca 1021 tcccaagcat catcagacca agccttaatc atcagattca acctcgatta agtcatgtgt 1081 tttaaatgca cctttggaga attgttcatc tcctcgacga attttttcca tgacgtaatt 1141 attagaaagt gttctcaacg tttcttgcat agaatcataa tctctttttg ataatacaac 1201 aactgtatct tctacatctt tacttgttac aataagtgtt tcagcatcct cattaacttg 1261 tttcatataa ctacgtaaat tttggcggaa atttgaataa gctactgctt ccattccttt 1321 caccccttat ttatttctat aaaacaattg tacattatat tgtacaatta agcaaatgat 1381 taatattcta acttcttata attaaaagtg ctatcacttt tagtttttag gataaaaggt 1441 gacgtcactt tttgagtgct ggtacctcaa atcctgcttt ttcttgggca atcaataaat 1501 ctaacgcaat acacataatc ctggattcac ttttgccgta cttctgcttt agttgctgca 1561 attctttttg tctcttttca tccaacgtaa cgggtaaccg tttatttcct attgcaattg 1621 gcattagtcg cttatggatc ctgaacttga ccgtaaacga atgccatatc aacgatttga 1681 gaggaaaaaa ggcaataatc atttataccg aaaagtgaga atgtgtgaaa taaggacaaa 1741 aaacctagta actaagggaa tggttcttat ttcgcaccaa tcgctccaac cgcccccgcc 1801 gcttaggtgg tgcgaaataa gcgggcaaca cccgataacc taagagaaag aaggagatcc 1861 atatgaataa aatcattcct ctattgtcag tcctatttgt tggcgaaata cttatgcgcc 1921 atgctgcagc tgtgcattta aacttcgtag tatttttggc aatactggta ttttgtttgg 1981 tcttttggcg ctttgtatta gcccttgtga gtctactatt caagatgatt gcagccatat 2041 ttttcatagg gttatttcga tgaaaggaga gagtatcgga tcccccgggc tcgaggtttc 2101 tagaagttgt ctcctcctgc actgactgac tgatacaatc gatttctgga tccgcaggcc 2161 tctgctagct tgactgactg agatacagcg taccttcagc tcacagacat gataagatac 2221 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa 2281 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac 2341 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc 2401 aagtaaaacc tctacaaatg tggtattggc ccatctctat cggtatcgta gcataacccc 2461 ttggggcctc taaacgggtc ttgaggggtt ttttgtgccc ctcgggccgg attgctatct 2521 accggcattg gcgcagaaaa aaatgcctga tgcgacgctg cgcgtcttat actcccacat 2581 atgccagatt cagcaacgga tacggcttcc ccaacttgcc cacttccata cgtgtcctcc 2641 ttaccagaaa tttatcctta aggtcgtcag ctatcctgca ggcgatctct cgatttcgat 2701 caagacattc ctttaatggt cttttctgga caccactagg ggtcagaagt agttcatcaa 2761 actttcttcc ctccctaatc tcattggtta ccttgggcta tcgaaactta attaaccagt 2821 caagtcagct acttggcgag atcgacttgt ctgggtttcg actacgctca gaattgcgtc 2881 agtcaagttc gatctggtcc ttgctattgc acccgttctc cgattacgag tttcatttaa 2941 atcatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg 3001 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg 3061 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg 3121 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga 3181 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc 3241 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt 3301 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact 3361 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg 3421 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt 3481 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt 3541 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct 3601 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg 3661 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt 3721 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt 3781 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcatt taaatttccg 3841 aactctccaa ggccctcgtc ggaaaatctt caaacctttc gtccgatcca tcttgcaggc 3901 tacctctcga acgaactatc gcaagtctct tggccggcct tgcgccttgg ctattgcttg 3961 gcagcgccta tcgccaggta ttactccaat cccgaatatc cgagatcggg atcacccgag 4021 agaagttcaa cctacatcct caatcccgat ctatccgaga tccgaggaat atcgaaatcg 4081 gggcgcgcca agctgttgtg accgcttgct ctagccagct atcgagttgt gaaccgatcc 4141 atctagcaat tggtctcgat ctagcgatag gcttcgatct agctatgtag aaacgccgtg 4201 tgctcgatcg cttgataagg tccacgtagc tgctataatt gcttcaacag aacatattga 4261 ctatccggta ttacccggca aaggaggtag ccaacatgac agctctgaca gagggagcga 4321 aactgttcga gaaggagatc ccgtacatca cagagttgga aggagatgtc gaagggatga 4381 aattcatcat caagggagaa ggaacgggtg acgcaacgac cggaacgatc aaggccaaat 4441 acatttgtac cacgggcgat ttgcctgtcc cctgggcgac gcttgtaagc acgctctcgt 4501 atggtgtcca gtgcttcgcg aaatatccat cgcacattaa ggactttttc aagtcggcca 4561 tgccagaagg ttacacacaa gaacgaacca tctcctttga gggggacgga gtgtataaga 4621 cacgcgcgat ggtaacgtac gagcgcgggt ccatctacaa cagggtaact cttactgggg 4681 agaactttaa gaaggacggg catatcttgc ggaaaaacgt ggcatttcaa tgtccgccct 4741 cgattctgta tattctcccg gacacggtga ataatgggat cagagtggag ttcaaccagg 4801 catacgatat tgagggtgtg actgaaaagc tcgtcaccaa atgcagccag atgaatcgcc 4861 cccttgcggg atcagcagcc gtccatatcc cccggtatca ccatattacc taccacacaa 4921 aactctcaaa agacagagat gagagaaggg accatatgtg cctggtcgaa gtagtgaagg 4981 cggtggatct tgatacatac cagtaggcta gccattggcg cagaaaaaaa tgcctgatgc 5041 gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac ggcttcccca 5101 acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaagg gcgcgcctgg 5161 tgtaccgaga acgatcctct cagtgcgagt ctcgacgatc catatcgttg cttggcagtc 5221 agccagtcgg aatccagctt gggacccagg aagtccaatc gtcagatatt gtactcaagc 5281 ctggtcacgg cagcgtaccg atctgtttaa acctagatat tgatagtctg atcggtcaac 5341 gtataatcga gtcctagctt ttgcaaacat ctatcaagag acaggatcag caggaggctt 5401 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gcggcttggg tggagaggct 5461 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct 5521 gtcagcgcag gggcgtccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga 5581 actgcaagac gaggcagcgc ggctatcgtg gctggcgacg acgggcgttc cttgcgcggc 5641 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg 5701 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc 5761 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca 5821 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga 5881 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgtctatgcc 5941 cgacggcgag gatctcgtcg tgacccacgg cgatgcctgc ttgccgaata tcatggtgga 6001 aaatggccgc ttttctggat tcatcgactg tggccgtctg ggtgtggcgg accgctatca 6061 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg 6121 cttccttgtg ctttacggta tcgccgcgcc cgattcgcag cgcatcgcct tctatcgcct 6181 tcttgacgag ttcttctgac cgattctagg tgcattggcg cagaaaaaaa tgcctgatgc 6241 gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac ggcttcccca 6301 acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaagg tcgtttaaac 6361 tcgactctgg ctctatcgaa tctccgtcgt ttcgagctta cgcgaacagc cgtggcgctc 6421 atttgctcgt cgggcatcga atctcgtcag ctatcgtcag cttacctttt tggca //