LOCUS       sequence-147449-        5726 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  LitR-L1 gate.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5726)
  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 5726)
  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: pLitR-L1
FEATURES             Location/Qualifiers
     source          1..5726
                     /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        2777..2826
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(2876..2878,2879..2881,2882..3595)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2921..2942)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3182..3201)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3529..3550
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3743..4825
                     /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(3762..3781)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4844..5467
                     /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(4917..4936)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(5650..5669)
                     /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 tagtcataag tctgggctaa gcccactgat gagtcgctga aatgcgacga
     1981 aacttatgac ctctacaaat aattttgttt aagtcctatg gactttttca tacaggagaa
     2041 ccctcgatgg ataccattca gaaacgtccg cgtacccgtc tgagtccgga aaaacgtaaa
     2101 gaacagctgc tggatattgc cattgaagtt tttagccagc gtggtattgg tcgtggtggt
     2161 catgcagata ttgcagaaat tgcacaggtt agcgttgcaa ccgtgtttaa ctattttccg
     2221 acccgtgaag atctggttga tgatgttctg aacaaagtgg aaaacgagtt tcaccagttc
     2281 atcaataaca gcattagcct ggatctggat gttcgtagca atctgaatac cctgctgctg
     2341 aacattattg atagcgttca gaccggcaac aaatggatta aagtttggtt tgaatggtca
     2401 accagcaccc gtgatgaagt ttggcctctg tttctgagca cccatagcaa taccaatcag
     2461 gtgatcaaaa ccatgtttga agagggtatt gaacgcaatg aagtgtgcaa tgatcataca
     2521 ccggaaaatc tgaccaaaat gctgcatggt atttgctata gcgtgtttat tcaggccaat
     2581 cgtaatagca gcagcgaaga aatggaagaa accgcaaatt gctttctgaa tatgctgtgc
     2641 atctacaaat aactcggtac caaattccag aaaagacacc cgaaagggtg ttttttcgtt
     2701 ttggtcctac tcgagcgtag agcttagatt cgttaccaat tgacaaattt ataaattgtc
     2761 agtacagtcc tagcagctgt caccggatgt gctttccggt ctgatgagtc cgtgaggacg
     2821 aaacagcctc tacaaataat tttgtttaat actagagaaa gaggggaaat actagatggt
     2881 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     2941 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     3001 gctgaccctg aagttcatct gcaccacagg caagctgccc gtgccctggc ccaccctcgt
     3061 gaccaccttc ggctacggcc tgcaatgctt cgcccgctac cccgaccaca tgaagctgca
     3121 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     3181 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     3241 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     3301 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     3361 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     3421 ctaccagcag aacaccccaa tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     3481 tagctaccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     3541 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtaactcgg
     3601 taccaaattc cagaaaagag gcctcccgaa aggggggcct tttttcgttt tggtccaatg
     3661 gcggcgcgcc atcgaatggc gcaaaacctt tcgcggtatg gcatgatagc gcccggaaga
     3721 gagtcaattc agggtggtga atatgaaacc agtaacgtta tacgatgtcg cagagtatgc
     3781 cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag gccagccacg tttctgcgaa
     3841 aacgcgggaa aaagtggaag cggcgatggc ggagctgaat tacattccca accgcgtggc
     3901 acaacaactg gcgggcaaac agtcgttgct gattggcgtt gccacctcca gtctggccct
     3961 gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac tgggtgccag
     4021 cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg cggtgcacaa
     4081 tcttctcgcg caacgcgtca gtgggctgat cattaactat ccgctggatg accaggatgc
     4141 cattgctgtg gaagctgcct gcactaatgt tccggcgtta tttcttgatg tctctgacca
     4201 gacacccatc aacagtatta ttttctccca tgaggacggt acgcgactgg gcgtggagca
     4261 tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa gttctgtctc
     4321 ggcgcgtctg cgtctggctg gctggcataa atatctcact cgcaatcaaa ttcagccgat
     4381 agcggaacgg gaaggcgact ggagtgccat gtccggtttt caacaaacca tgcaaatgct
     4441 gaatgagggc atcgttccca ctgcgatgct ggttgccaac gatcagatgg cgctgggcgc
     4501 aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg gatatctcgg tagtgggata
     4561 cgacgatacc gaagatagct catgttatat cccgccgtta accaccatca aacaggattt
     4621 tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg gccaggcggt
     4681 gaagggcaat cagctgttgc cagtctcact ggtgaaaaga aaaaccaccc tggcgcccaa
     4741 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
     4801 ttcccgactg gaaagcgggc agtgataatc caggaggaaa aaaatgtcca gattagataa
     4861 aagtaaagtg attaacagcg cattagagct gcttaatgag gtcggaatcg aaggtttaac
     4921 aacccgtaaa ctcgcccaga agctaggtgt agagcagcct acattgtatt ggcatgtaaa
     4981 aaataagcgg gctttgctcg acgccttagc cattgagatg ttagataggc accatactca
     5041 cttttgccct ttagaagggg aaagctggca agatttttta cgtaataacg ctaaaagttt
     5101 tagatgtgct ttactaagtc atcgcgatgg agcaaaagta catttaggta cacggcctac
     5161 agaaaaacag tatgaaactc tcgaaaatca attagccttt ttatgccaac aaggtttttc
     5221 actagagaat gcattatatg cactcagcgc tgtggggcat tttactttag gttgcgtatt
     5281 ggaagatcaa gagcatcaag tcgctaaaga agaaagggaa acacctacta ctgatagtat
     5341 gccgccatta ttacgacaag ctatcgaatt atttgatcac caaggtgcag agccagcctt
     5401 cttattcggc cttgaattga tcatatgcgg attagaaaaa caacttaaat gtgaaagtgg
     5461 gtcctaataa ttggtaacga atcagacaat tgacggctcg agggagtagc atagggtttg
     5521 cagaatccct gcttcgtcca tttgacaggc acattatgca tcgatgataa gctgtcaaac
     5581 atgagcagat cctctacgcc ggacgcatcg tggccggcat caccggcgcc acaggtgcgg
     5641 ttgctggcgc ctatatcgcc gacatcaccg atggggaaga tcgggctcgc cacttcgggc
     5701 tcatgagcaa atattttatc tgaggt
//
