LOCUS       sequence-141098-        5767 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AmeR-F1 gate.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5767)
  AUTHORS   Nielsen AA, Der BS, Shin J, Vaidyanathan P, Paralanov V, Strychalski
            EA, Ross D, Densmore D, Voigt CA
  TITLE     Genetic circuit design automation.
  JOURNAL   Science. 2016 Apr 1;352(6281):aac7341. doi: 10.1126/science.aac7341.
  PUBMED    27034378
REFERENCE   2  (bases 1 to 5767)
  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: "Science."; 
            date: "2016-04-1"; volume: "352(6281):aac7341. doi"; pages: " 
            10.1126/science.aac7341"
COMMENT     Sequence Label: pAmeR-F1
FEATURES             Location/Qualifiers
     source          1..5767
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(308..852)
                     /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(388..407)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(941..1756)
                     /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(941..1756)
                     /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     1664..1683
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1748..1767
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     promoter        join(1878..1883,1884..1899,1900..1906)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1912..1931
                     /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."
     misc_RNA        2818..2867
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(2917..2919,2920..2922,2923..3636)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2962..2983)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3223..3242)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3570..3591
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3784..4866
                     /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(3803..3822)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4885..5508
                     /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(4958..4977)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(5691..5710)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 gcttcctcgc tcactgactc gctgcacgag gcagacctca gcgctagcgg agtgtatact
       61 ggcttactat gttggcactg atgagggtgt cagtgaagtg cttcatgtgg caggagaaaa
      121 aaggctgcac cggtgcgtca gcagaatatg tgatacagga tatattccgc ttcctcgctc
      181 actgactcgc tacgctcggt cgttcgactg cggcgagcgg aaatggctta cgaacggggc
      241 ggagatttcc tggaagatgc caggaagata cttaacaggg aagtgagagg gccgcggcaa
      301 agccgttttt ccataggctc cgcccccctg acaagcatca cgaaatctga cgctcaaatc
      361 agtggtggcg aaacccgaca ggactataaa gataccaggc gtttcccctg gcggctccct
      421 cgtgcgctct cctgttcctg cctttcggtt taccggtgtc attccgctgt tatggccgcg
      481 tttgtctcat tccacgcctg acactcagtt ccgggtaggc agttcgctcc aagctggact
      541 gtatgcacga accccccgtt cagtccgacc gctgcgcctt atccggtaac tatcgtcttg
      601 agtccaaccc ggaaagacat gcaaaagcac cactggcagc agccactggt aattgattta
      661 gaggagttag tcttgaagtc atgcgccggt taaggctaaa ctgaaaggac aagttttggt
      721 gactgcgctc ctccaagcca gttacctcgg ttcaaagagt tggtagctca gagaaccttc
      781 gaaaaaccgc cctgcaaggc ggttttttcg ttttcagagc aagagattac gcgcagacca
      841 aaacgatctc aagaagatca tcttattaag gggtctgacg ctcagtggaa cgaaaaatca
      901 atctaaagta tatatgagta aacttggtct gacagttacc ttagaaaaac tcatcgagca
      961 tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc
     1021 gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt
     1081 atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa
     1141 aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca
     1201 aaagcttatg catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa
     1261 aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata
     1321 cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca
     1381 ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg
     1441 ctgttttccc ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat
     1501 gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg
     1561 taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct
     1621 tcccatacaa tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat
     1681 acccatataa atcagcatcc atgttggaat ttaatcgcgg cctcgagcaa gacgtttccc
     1741 gttgaatatg gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg
     1801 ttcatgatga tatattttta tcttgtgcaa tgtacatcag agattttgag acacaaggag
     1861 aacgatcgtt ggctgtgttg acaattaatc atcggctcgt ataatgtgtg gaattgtgag
     1921 cgctcacaat taggggtcag ttgatgtgct ttcaactctg atgagtcagt gatgacgaaa
     1981 ccccctctac aaataatttt gtttaactat ggactatgtt ttcacatacg agggggatta
     2041 gatgaacaaa accattgatc aggtgcgtaa aggtgatcgt aaaagcgatc tgccggttcg
     2101 tcgtcgtccg cgtcgtagtg ccgaagaaac ccgtcgtgat attctggcaa aagccgaaga
     2161 actgtttcgt gaacgtggtt ttaatgcagt tgccattgca gatattgcaa gcgcactgaa
     2221 tatgagtccg gcaaatgtgt ttaaacattt tagcagcaaa aacgcactgg ttgatgcaat
     2281 tggttttggt cagattggtg tttttgaacg tcagatttgt ccgctggata aaagccatgc
     2341 accgctggat cgtctgcgtc atctggcacg taatctgatg gaacagcatc atcaggatca
     2401 tttcaaacac atacgggttt ttattcagat cctgatgacc gccaaacagg atatgaaatg
     2461 tggcgattat tacaaaagcg tgattgcaaa actgctggcc gaaattattc gtgatggtgt
     2521 tgaagcaggt ctgtatattg caaccgatat tccggttctg gcagaaaccg ttctgcatgc
     2581 actgaccagc gttattcatc cggttctgat tgcacaagaa gatattggta atctggcaac
     2641 ccgttgtgat cagctggttg atctgattga tgcaggtctg cgtaatccgc tggcaaaata
     2701 accaattatt gaacacccta acgggtgttt tttttttttt ggtctcccta cttcgtcact
     2761 agagggcgat agtgacaaac ttgacaactc atcacttcct aggtataatg ctagcagctg
     2821 tcaccggatg tgctttccgg tctgatgagt ccgtgaggac gaaacagcct ctacaaataa
     2881 ttttgtttaa tactagagaa agaggggaaa tactagatgg tgagcaaggg cgaggagctg
     2941 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
     3001 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
     3061 tgcaccacag gcaagctgcc cgtgccctgg cccaccctcg tgaccacctt cggctacggc
     3121 ctgcaatgct tcgcccgcta ccccgaccac atgaagctgc acgacttctt caagtccgcc
     3181 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
     3241 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
     3301 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc
     3361 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc
     3421 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacacccca
     3481 atcggcgacg gccccgtgct gctgcccgac aaccactacc ttagctacca gtccgccctg
     3541 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     3601 gggatcactc tcggcatgga cgagctgtac aagtaactcg gtaccaaatt ccagaaaaga
     3661 ggcctcccga aaggggggcc ttttttcgtt ttggtccaat ggcggcgcgc catcgaatgg
     3721 cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag agagtcaatt cagggtggtg
     3781 aatatgaaac cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc ttatcagacc
     3841 gtttcccgcg tggtgaacca ggccagccac gtttctgcga aaacgcggga aaaagtggaa
     3901 gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg cacaacaact ggcgggcaaa
     3961 cagtcgttgc tgattggcgt tgccacctcc agtctggccc tgcacgcgcc gtcgcaaatt
     4021 gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt gtcgatggta
     4081 gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc gcaacgcgtc
     4141 agtgggctga tcattaacta tccgctggat gaccaggatg ccattgctgt ggaagctgcc
     4201 tgcactaatg ttccggcgtt atttcttgat gtctctgacc agacacccat caacagtatt
     4261 attttctccc atgaggacgg tacgcgactg ggcgtggagc atctggtcgc attgggtcac
     4321 cagcaaatcg cgctgttagc gggcccatta agttctgtct cggcgcgtct gcgtctggct
     4381 ggctggcata aatatctcac tcgcaatcaa attcagccga tagcggaacg ggaaggcgac
     4441 tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc
     4501 actgcgatgc tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc cattaccgag
     4561 tccgggctgc gcgttggtgc ggatatctcg gtagtgggat acgacgatac cgaagatagc
     4621 tcatgttata tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc
     4681 agcgtggacc gcttgctgca actctctcag ggccaggcgg tgaagggcaa tcagctgttg
     4741 ccagtctcac tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac cgcctctccc
     4801 cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg
     4861 cagtgataat ccaggaggaa aaaaatgtcc agattagata aaagtaaagt gattaacagc
     4921 gcattagagc tgcttaatga ggtcggaatc gaaggtttaa caacccgtaa actcgcccag
     4981 aagctaggtg tagagcagcc tacattgtat tggcatgtaa aaaataagcg ggctttgctc
     5041 gacgccttag ccattgagat gttagatagg caccatactc acttttgccc tttagaaggg
     5101 gaaagctggc aagatttttt acgtaataac gctaaaagtt ttagatgtgc tttactaagt
     5161 catcgcgatg gagcaaaagt acatttaggt acacggccta cagaaaaaca gtatgaaact
     5221 ctcgaaaatc aattagcctt tttatgccaa caaggttttt cactagagaa tgcattatat
     5281 gcactcagcg ctgtggggca ttttacttta ggttgcgtat tggaagatca agagcatcaa
     5341 gtcgctaaag aagaaaggga aacacctact actgatagta tgccgccatt attacgacaa
     5401 gctatcgaat tatttgatca ccaaggtgca gagccagcct tcttattcgg ccttgaattg
     5461 atcatatgcg gattagaaaa acaacttaaa tgtgaaagtg ggtcctaata attggtaacg
     5521 aatcagacaa ttgacggctc gagggagtag catagggttt gcagaatccc tgcttcgtcc
     5581 atttgacagg cacattatgc atcgatgata agctgtcaaa catgagcaga tcctctacgc
     5641 cggacgcatc gtggccggca tcaccggcgc cacaggtgcg gttgctggcg cctatatcgc
     5701 cgacatcacc gatggggaag atcgggctcg ccacttcggg ctcatgagca aatattttat
     5761 ctgaggt
//
