LOCUS       sequence-272327-        7430 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tunable NOR gate.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7430)
  AUTHORS   Bartoli V, Meaker GA, di Bernardo M, Gorochowski TE
  TITLE     Tunable genetic devices through simultaneous control of 
            transcription and translation.
  JOURNAL   Nat Commun. 2020 Apr 29;11(1):2095. doi: 10.1038/s41467-020-15653-7.
  PUBMED    32350250
REFERENCE   2  (bases 1 to 7430)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2020-04-29"; volume: "11(1):2095. doi"; pages: " 
            10.1038/s41467-020-15653-7"
COMMENT     Sequence Label: pVB006
FEATURES             Location/Qualifiers
     source          1..7430
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(130..945)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(130..945)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     853..872
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     937..956
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     promoter        join(1146..1151,1152..1167,1168..1174)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1180..1199
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     promoter        1338..1622
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     1545..1564
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     protein_bind    1663..1681
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1688..1706
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_RNA        1719..1768
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     misc_RNA        2669..2718
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(2768..2770,2771..2773,2774..3487)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2813..2834)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3074..3093)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3421..3442
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        3597..3631
                     /label=BBa_J23105
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.24"
     RBS             3640..3651
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             4507..4518
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             4636..5718
                     /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(4655..4674)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             5737..6360
                     /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(5810..5829)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(6543..6562)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     rep_origin      complement(join(6927..7430,1..41))
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(7007..7026)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tttcagagca agagattacg cgcagaccaa aacgatctca agaagatcat cttattaagg
       61 ggtctgacgc tcagtggaac gaaaaatcaa tctaaagtat atatgagtaa acttggtctg
      121 acagttacct tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg
      181 attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag
      241 gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc
      301 aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg
      361 agtgacgact gaatccggtg agaatggcaa aagcttatgc atttctttcc agacttgttc
      421 aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat
      481 tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac
      541 aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga
      601 atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag tggtgagtaa
      661 ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt
      721 cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg
      781 tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg tcgcacctga
      841 ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt
      901 taatcgcggc ctcgagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt
      961 actgtttatg taagcagaca gttttattgt tcatgatgat atatttttat cttgtgcaat
     1021 gtacatcaga gattttgaga cacaaccaat tattgaaggc ctccctaacg gggggccttt
     1081 ttttgtttct ggtctcccgc ttacttttca tactcccgcc gcggccgcaa cgatcgttgg
     1141 ctgtgttgac aattaatcat cggctcgtat aatgtgtgga attgtgagcg ctcacaattg
     1201 ggtcatgact gggacacgcc agtcatgaga atacagacat aaagcaagcc agagattaac
     1261 gaagccaatt attgaacacc cttcggggtg tttttttgtt tctggtctac cacttttcat
     1321 actcccgcca ttcagagaag aaaccaattg tccatattgc atcagacatt gccgtcactg
     1381 cgtcttttac tggctcttct cgctaaccaa accggtaacc ccgcttatta aaagcattct
     1441 gtaacaaagc gggaccaaag ccatgacaaa aacgcgtaac aaaagtgtct ataatcacgg
     1501 cagaaaagtc cacattgatt atttgcacgg cgtcacactt tgctatgcca tagcattttt
     1561 atccataaga ttagcggatc ctacctgacg ctttttatcg caactctcta ctgtttctcc
     1621 atacccgttt ttttgggcta gctactccac cgttggcttt tttccctatc agtgatagag
     1681 attgacatcc ctatcagtga tagagataat gagcacagct gtcaccggat gtgctttccg
     1741 gtctgatgag tccgtgagga cgaaacagcc tctacaaata attttgttta agggcgttaa
     1801 tctctggctt gctttatgtc tgtaaacaga ggagatacag aatgaaagca agcaacctgg
     1861 cggcagcgca aaagatgcgt aaagcacgta ccccgagccg tagcagcatt ggtagcctgc
     1921 gtagtccgca tacccataaa gcaattctga ccagcaccat tgaaatcctg aaagaatgtg
     1981 gttatagcgg tctgagcatt gaaagcgttg cacgtcgtgc cggtgcaagc aaaccgacca
     2041 tttatcgttg gtggaccaat aaagcagcac tgattgccga agtgtatgaa aatgaaagcg
     2101 aacaggtgcg taaatttccg gatctgggta gctttaaagc cgatctggat tttctgctgc
     2161 gtaatctgtg gaaagtttgg cgtgaaacca tttgtggtga agcatttcgt tgtgttattg
     2221 cagaagcaca gctggaccct gcaaccctga cccagctgaa agatcagttt atggaacgtc
     2281 gtcgtgagat gccgaaaaaa ctggttgaaa atgccattag caatggtgaa ctgccgaaag
     2341 ataccaatcg tgaactgctg ctggatatga tttttggttt ttgttggtat cgcctgctga
     2401 ccgaacagct gaccgttgaa caggatattg aagaatttac cttcctgctg attaatggtg
     2461 tttgtccggg tacacagcgt taaggaaaca cagaaaaaag cccgcacctg acagtgcggg
     2521 cttcgatcaa ctcgcgccgc tggtcttacg gtctcatgac agtgcgggct ttttttttcg
     2581 accaaaggtg tcaacgttcg acgtacggtg gaatctgatt cgttaccaat tgacatgata
     2641 cgaaacgtac cgtatcgtta aggtcaagct gtcaccggat gtgctttccg gtctgatgag
     2701 tccgtgagga cgaaacagcc tctacaaata attttgttta atactagaga aagaggggaa
     2761 atactagatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     2821 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     2881 cacctacggc aagctgaccc tgaagttcat ctgcaccaca ggcaagctgc ccgtgccctg
     2941 gcccaccctc gtgaccacct tcggctacgg cctgcaatgc ttcgcccgct accccgacca
     3001 catgaagctg cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     3061 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     3121 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     3181 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca
     3241 gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     3301 gctcgccgac cactaccagc agaacacccc aatcggcgac ggccccgtgc tgctgcccga
     3361 caaccactac cttagctacc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     3421 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     3481 caagtaagaa ttccatggac gagctgtaca agtaactcgg taccaaattc cagaaaagag
     3541 gcctcccgaa aggggggcct tttttcgttt tggtccaatg gtcaccatat atcaagttta
     3601 cggctagctc agtcctaggt actatgctag ctactagaga aagaggagaa atactagatg
     3661 gctgaagcgc aaaatgatcc cctgctgccg ggatactcgt ttaatgccca tctggtggcg
     3721 ggtttaacgc cgattgaggc caacggttat ctcgattttt ttatcgaccg accgctggga
     3781 atgaaaggtt atattctcaa tctcaccatt cgcggtcagg gggtggtgaa aaatcaggga
     3841 cgagaatttg tttgccgacc gggtgatatt ttgctgttcc cgccaggaga gattcatcac
     3901 tacggtcgtc atccggaggc tcgcgaatgg tatcaccagt gggtttactt tcgtccgcgc
     3961 gcctactggc atgaatggct taactggccg tcaatatttg ccaatacggg gttctttcgc
     4021 ccggatgaag cgcaccagcc gcatttcagc gacctgtttg ggcaaatcat taacgccggg
     4081 caaggggaag ggcgctattc ggagctgctg gcgataaatc tgcttgagca attgttactg
     4141 cggcgcatgg aagcgattaa cgagtcgctc catccaccga tggataatcg ggtacgcgag
     4201 gcttgtcagt acatcagcga tcacctggca gacagcaatt ttgatatcgc cagcgtcgca
     4261 cagcatgttt gcttgtcgcc gtcgcgtctg tcacatcttt tccgccagca gttagggatt
     4321 agcgtcttaa gctggcgcga ggaccaacgt atcagccagg cgaagctgct tttgagcacc
     4381 acccggatgc ctatcgccac cgtcggtcgc aatgttggtt ttgacgatca actctatttc
     4441 tcgcgggtat ttaaaaaatg caccggggcc agcccgagcg agttccgtgc cggttaataa
     4501 ccaattattg aaggccgcta acgcagcctt tttttgtttc tggtctccca atggcggcgc
     4561 gccatcgaat ggcgcaaaac ctttcgcggt atggcatgat agcgcccgga agagagtcaa
     4621 ttcagggtgg tgaatatgaa accagtaacg ttatacgatg tcgcagagta tgccggtgtc
     4681 tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc gaaaacgcgg
     4741 gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt ggcacaacaa
     4801 ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc cctgcacgcg
     4861 ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc cagcgtggtg
     4921 gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca caatcttctc
     4981 gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga tgccattgct
     5041 gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga ccagacaccc
     5101 atcaacagta ttattttctc ccatgaggac ggtacgcgac tgggcgtgga gcatctggtc
     5161 gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt ctcggcgcgt
     5221 ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc gatagcggaa
     5281 cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat gctgaatgag
     5341 ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg cgcaatgcgc
     5401 gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg atacgacgat
     5461 accgaagata gctcatgtta tatcccgccg ttaaccacca tcaaacagga ttttcgcctg
     5521 ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc ggtgaagggc
     5581 aatcagctgt tgccagtctc actggtgaaa agaaaaacca ccctggcgcc caatacgcaa
     5641 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     5701 ctggaaagcg ggcagtgata atccaggagg aaaaaaatgt ccagattaga taaaagtaaa
     5761 gtgattaaca gcgcattaga gctgcttaat gaggtcggaa tcgaaggttt aacaacccgt
     5821 aaactcgccc agaagctagg tgtagagcag cctacattgt attggcatgt aaaaaataag
     5881 cgggctttgc tcgacgcctt agccattgag atgttagata ggcaccatac tcacttttgc
     5941 cctttagaag gggaaagctg gcaagatttt ttacgtaata acgctaaaag ttttagatgt
     6001 gctttactaa gtcatcgcga tggagcaaaa gtacatttag gtacacggcc tacagaaaaa
     6061 cagtatgaaa ctctcgaaaa tcaattagcc tttttatgcc aacaaggttt ttcactagag
     6121 aatgcattat atgcactcag cgctgtgggg cattttactt taggttgcgt attggaagat
     6181 caagagcatc aagtcgctaa agaagaaagg gaaacaccta ctactgatag tatgccgcca
     6241 ttattacgac aagctatcga attatttgat caccaaggtg cagagccagc cttcttattc
     6301 ggccttgaat tgatcatatg cggattagaa aaacaactta aatgtgaaag tgggtcctaa
     6361 taattggtaa cgaatcagac aattgacggc tcgagggagt agcatagggt ttgcagaatc
     6421 cctgcttcgt ccatttgaca ggcacattat gcatcgatga taagctgtca aacatgagca
     6481 gatcctctac gccggacgca tcgtggccgg catcaccggc gccacaggtg cggttgctgg
     6541 cgcctatatc gccgacatca ccgatgggga agatcgggct cgccacttcg ggctcatgag
     6601 caaatatttt atctgaggtg cttcctcgct cactgactcg ctgcacgagg cagacctcag
     6661 cgctagcgga gtgtatactg gcttactatg ttggcactga tgagggtgtc agtgaagtgc
     6721 ttcatgtggc aggagaaaaa aggctgcacc ggtgcgtcag cagaatatgt gatacaggat
     6781 atattccgct tcctcgctca ctgactcgct acgctcggtc gttcgactgc ggcgagcgga
     6841 aatggcttac gaacggggcg gagatttcct ggaagatgcc aggaagatac ttaacaggga
     6901 agtgagaggg ccgcggcaaa gccgtttttc cataggctcc gcccccctga caagcatcac
     6961 gaaatctgac gctcaaatca gtggtggcga aacccgacag gactataaag ataccaggcg
     7021 tttcccctgg cggctccctc gtgcgctctc ctgttcctgc ctttcggttt accggtgtca
     7081 ttccgctgtt atggccgcgt ttgtctcatt ccacgcctga cactcagttc cgggtaggca
     7141 gttcgctcca agctggactg tatgcacgaa ccccccgttc agtccgaccg ctgcgcctta
     7201 tccggtaact atcgtcttga gtccaacccg gaaagacatg caaaagcacc actggcagca
     7261 gccactggta attgatttag aggagttagt cttgaagtca tgcgccggtt aaggctaaac
     7321 tgaaaggaca agttttggtg actgcgctcc tccaagccag ttacctcggt tcaaagagtt
     7381 ggtagctcag agaaccttcg aaaaaccgcc ctgcaaggcg gttttttcgt
//
