LOCUS       sequence-141196-        5814 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SrpR-S2 gate.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5814)
  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 5814)
  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: pSrpR-S2
FEATURES             Location/Qualifiers
     source          1..5814
                     /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        2865..2914
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(2964..2966,2967..2969,2970..3683)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3009..3030)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3270..3289)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3617..3638
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3831..4913
                     /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(3850..3869)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4932..5555
                     /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(5005..5024)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(5738..5757)
                     /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 tagcgctcaa cgggtgtgct tcccgttctg atgagtccgt gaggacgaaa
     1981 gcgcctctac aaataatttt gtttaagagt ctatggacta tgttttcaca tatgagatac
     2041 caggatggca cgtaaaaccg cagcagaagc agaagaaacc cgtcagcgta ttattgatgc
     2101 agcactggaa gtttttgttg cacagggtgt tagtgatgca accctggatc agattgcacg
     2161 taaagccggt gttacccgtg gtgcagttta ttggcatttt aatggtaaac tggaagttct
     2221 gcaggcagtt ctggcaagcc gtcagcatcc gctggaactg gattttacac cggatctggg
     2281 tattgaacgt agctgggaag cagttgttgt tgcaatgctg gatgcagttc atagtccgca
     2341 gagcaaacag tttagcgaaa ttctgattta tcagggtctg gatgaaagcg gtctgattca
     2401 taatcgtatg gttcaggcaa gcgatcgttt tctgcagtat attcatcagg ttctgcgtca
     2461 tgcagttacc cagggtgaac tgccgattaa tctggatctg cagaccagca ttggtgtttt
     2521 taaaggtctg attaccggtc tgctgtatga aggtctgcgt agcaaagatc agcaggcaca
     2581 gattatcaaa gttgcactgg gtagcttttg ggcactgctg cgtgaaccgc ctcgttttct
     2641 gctgtgtgaa gaagcacaga ttaaacaggt gaaatccttc gaataattca gccaaaaaac
     2701 ttaagaccgc cggtcttgtc cactaccttg cagtaatgcg gtggacagga tcggcggttt
     2761 tcttttctct tctcaatact tctatgattg gtccagattc gttaccaatt gacagctagc
     2821 tcagtcctag gtatatacat acatgcttgt ttgtttgtaa acagctgtca ccggatgtgc
     2881 tttccggtct gatgagtccg tgaggacgaa acagcctcta caaataattt tgtttaatac
     2941 tagagaaaga ggggaaatac tagatggtga gcaagggcga ggagctgttc accggggtgg
     3001 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     3061 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accacaggca
     3121 agctgcccgt gccctggccc accctcgtga ccaccttcgg ctacggcctg caatgcttcg
     3181 cccgctaccc cgaccacatg aagctgcacg acttcttcaa gtccgccatg cccgaaggct
     3241 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     3301 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     3361 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     3421 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg
     3481 aggacggcag cgtgcagctc gccgaccact accagcagaa caccccaatc ggcgacggcc
     3541 ccgtgctgct gcccgacaac cactacctta gctaccagtc cgccctgagc aaagacccca
     3601 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     3661 gcatggacga gctgtacaag taactcggta ccaaattcca gaaaagaggc ctcccgaaag
     3721 gggggccttt tttcgttttg gtccaatggc ggcgcgccat cgaatggcgc aaaacctttc
     3781 gcggtatggc atgatagcgc ccggaagaga gtcaattcag ggtggtgaat atgaaaccag
     3841 taacgttata cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg
     3901 tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg
     3961 agctgaatta cattcccaac cgcgtggcac aacaactggc gggcaaacag tcgttgctga
     4021 ttggcgttgc cacctccagt ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta
     4081 aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg
     4141 tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca
     4201 ttaactatcc gctggatgac caggatgcca ttgctgtgga agctgcctgc actaatgttc
     4261 cggcgttatt tcttgatgtc tctgaccaga cacccatcaa cagtattatt ttctcccatg
     4321 aggacggtac gcgactgggc gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc
     4381 tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg tctggctggc tggcataaat
     4441 atctcactcg caatcaaatt cagccgatag cggaacggga aggcgactgg agtgccatgt
     4501 ccggttttca acaaaccatg caaatgctga atgagggcat cgttcccact gcgatgctgg
     4561 ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg
     4621 ttggtgcgga tatctcggta gtgggatacg acgataccga agatagctca tgttatatcc
     4681 cgccgttaac caccatcaaa caggattttc gcctgctggg gcaaaccagc gtggaccgct
     4741 tgctgcaact ctctcagggc caggcggtga agggcaatca gctgttgcca gtctcactgg
     4801 tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg
     4861 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgataatcca
     4921 ggaggaaaaa aatgtccaga ttagataaaa gtaaagtgat taacagcgca ttagagctgc
     4981 ttaatgaggt cggaatcgaa ggtttaacaa cccgtaaact cgcccagaag ctaggtgtag
     5041 agcagcctac attgtattgg catgtaaaaa ataagcgggc tttgctcgac gccttagcca
     5101 ttgagatgtt agataggcac catactcact tttgcccttt agaaggggaa agctggcaag
     5161 attttttacg taataacgct aaaagtttta gatgtgcttt actaagtcat cgcgatggag
     5221 caaaagtaca tttaggtaca cggcctacag aaaaacagta tgaaactctc gaaaatcaat
     5281 tagccttttt atgccaacaa ggtttttcac tagagaatgc attatatgca ctcagcgctg
     5341 tggggcattt tactttaggt tgcgtattgg aagatcaaga gcatcaagtc gctaaagaag
     5401 aaagggaaac acctactact gatagtatgc cgccattatt acgacaagct atcgaattat
     5461 ttgatcacca aggtgcagag ccagccttct tattcggcct tgaattgatc atatgcggat
     5521 tagaaaaaca acttaaatgt gaaagtgggt cctaataatt ggtaacgaat cagacaattg
     5581 acggctcgag ggagtagcat agggtttgca gaatccctgc ttcgtccatt tgacaggcac
     5641 attatgcatc gatgataagc tgtcaaacat gagcagatcc tctacgccgg acgcatcgtg
     5701 gccggcatca ccggcgccac aggtgcggtt gctggcgcct atatcgccga catcaccgat
     5761 ggggaagatc gggctcgcca cttcgggctc atgagcaaat attttatctg aggt
//
