LOCUS       sequence-108253-        7197 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic autoregulatory gene circuit made by inserting an intron 
            containing the mouse mir-124–3 gene into mCherry. Does not contain 
            the target containing Vamp3 3′UTR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7197)
  AUTHORS   Strovas TJ, Rosenberg AB, Kuypers BE, Muscat RA, Seelig G
  TITLE     MicroRNA-based single-gene circuits buffer protein synthesis rates 
            against perturbations.
  JOURNAL   ACS Synth Biol. 2014 May 16;3(5):324-31. doi: 10.1021/sb4001867. 
            Epub 2014 Jan 30.
  PUBMED    24847681
REFERENCE   2  (bases 1 to 7197)
  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"; doi: "10.1021/sb4001867";
            journalName: "ACS Synth Biol"; date: "2014-05-16- 16"; volume: "3"; 
            issue: "5"; pages: "324-31"
COMMENT     Sequence Label: OpenLoopControl
FEATURES             Location/Qualifiers
     source          1..7197
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             992..1369
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     regulatory      1388..1397
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1539..1557)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1807..1826
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             complement(2642..2653)
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     primer_bind     complement(3149..3166)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3155..3379
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3512..3531)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3663..3710
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             3718..4740
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3718..4740
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4870..4991
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4907..4926)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4961..4980
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5040..5056)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5040..5056)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5053..5075)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5064..5080
                     /label=lac operator
                     /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)."
     promoter        complement(join(5088..5094,5095..5112,5113..5118))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5133..5154
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5271..5288)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5442..6030)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5522..5541)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6201..6992,6993..7061))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6824..6843
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7062..7166)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag ctttcgccac catgccagag ccagcgaagt ctgctcccgc
     1021 cccgaaaaag ggctccaaga aggcggtgac taaggcgcag aagaaaggcg gcaagaagcg
     1081 caagcgcagc cgcaaggaga gctattccat ctatgtgtac aaggttctga agcaggtcca
     1141 ccctgacacc ggcatttcgt ccaaggccat gggcatcatg aattcgtttg tgaacgacat
     1201 tttcgagcgc atcgcaggtg aggcttcccg cctggcgcat tacaacaagc gctcgaccat
     1261 cacctccagg gagatccaga cggccgtgcg cctgctgctg cctggggagt tggccaagca
     1321 cgccgtgtcc gagggtacta aggccatcac caagtacacc agcgctaagg atccccgggt
     1381 accggtcgcc accatggtga gcaagggcga ggaggataac atggccatca tcaaggagtt
     1441 catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg
     1501 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
     1561 tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc
     1621 ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt
     1681 caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc
     1741 ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc
     1801 cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta
     1861 ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg
     1921 ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc aggtgagtac
     1981 tccctctcaa aagcgggcat gacttctgcg ctaagattgt cagtttccaa aaacgaggag
     2041 gatttgatat tcacctggcc cgcggtgatg cctttgaggg tggccgcgtc catctggtca
     2101 gaaaagacaa tctttttgtt gtcaagctgt ttgactcatg cacccgctac tcttcaacct
     2161 tgcagactaa ttaatggcaa ctctgacaga tgtggcggga ggaaagctga ggtgggagta
     2221 ctgctcagag ctacaactct aggagtaggg actccaagcc tagagctcca agagagggtg
     2281 aagggcaggg agaaaattat agtaatagtt gcaatgagtc acttgcttct agatcaagat
     2341 cagagactct gctctccgtg ttcacagcgg accttgattt aatgtcatac aattaaggca
     2401 cgcggtgaat gccaagagcg gagcctacgg ctgcacttga aggacatccg agagaagtta
     2461 ggaagggtgg ggagaaacaa ttctagaatg aacccatcct gtgcgacaca aggacccctc
     2521 gcattgattc aaaatcttta gaatacaaga cagtcaaatg tttttcagcg agaatctggg
     2581 aaatccgcat gcatactccc aaggggaata ttagaggttt cattgacagc tgtcaggaga
     2641 gctgtagttc ccattggtgc aaacggtcta agagatggca ctcagagagt gctggccttt
     2701 tccaaatact agtcatatgc ggccgcagct tgaggtgtga caggcttgag atctggccat
     2761 acacttgagt gacaatgaca tccactttgc ctttctctcc acagctgccc ggcgcctaca
     2821 acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc gtggaacagt
     2881 acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac aagtctgacc
     2941 ttgatatcca gcacagtggc ggccgctcga gtagccatgg cttcccgccg gcggtggcgg
     3001 cgcaggatga tggcacgctg cccatgtctt gtgcccagga gagcgggatg gaccgtcacc
     3061 ctgcagcctg tgcttctgct aggatcaatg tgtagtaatg agtttaaggc gcgcggctct
     3121 agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     3181 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     3241 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     3301 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     3361 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     3421 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3481 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     3541 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     3601 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtacc
     3661 tagaagttcc tattccgaag ttcctattct ctagaaagta taggaacttc cttggccaaa
     3721 aagcctgaac tcaccgcgac gtctgtcgag aagtttctga tcgaaaagtt cgacagcgtc
     3781 tccgacctga tgcagctctc ggagggcgaa gaatctcgtg ctttcagctt cgatgtagga
     3841 gggcgtggat atgtcctgcg ggtaaatagc tgcgccgatg gtttctacaa agatcgttat
     3901 gtttatcggc actttgcatc ggccgcgctc ccgattccgg aagtgcttga cattggggaa
     3961 ttcagcgaga gcctgaccta ttgcatctcc cgccgtgcac agggtgtcac gttgcaagac
     4021 ctgcctgaaa ccgaactgcc cgctgttctg cagccggtcg cggaggccat ggatgcgatc
     4081 gctgcggccg atcttagcca gacgagcggg ttcggcccat tcggaccgca aggaatcggt
     4141 caatacacta catggcgtga tttcatatgc gcgattgctg atccccatgt gtatcactgg
     4201 caaactgtga tggacgacac cgtcagtgcg tccgtcgcgc aggctctcga tgagctgatg
     4261 ctttgggccg aggactgccc cgaagtccgg cacctcgtgc acgcggattt cggctccaac
     4321 aatgtcctga cggacaatgg ccgcataaca gcggtcattg actggagcga ggcgatgttc
     4381 ggggattccc aatacgaggt cgccaacatc ttcttctgga ggccgtggtt ggcttgtatg
     4441 gagcagcaga cgcgctactt cgagcggagg catccggagc ttgcaggatc gccgcggctc
     4501 cgggcgtata tgctccgcat tggtcttgac caactctatc agagcttggt tgacggcaat
     4561 ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc cggagccggg
     4621 actgtcgggc gtacacaaat cgcccgcaga agcgcggccg tctggaccga tggctgtgta
     4681 gaagtactcg ccgatagtgg aaaccgacgc cccagcactc gtccgagggc aaaggaatag
     4741 cacgtactac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     4801 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     4861 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     4921 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     4981 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     5041 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     5101 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     5161 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     5221 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     5281 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5341 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5401 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5461 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5521 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5581 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5641 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     5701 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5761 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5821 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5881 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5941 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     6001 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     6061 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     6121 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     6181 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     6241 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6301 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6361 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6421 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6481 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6541 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6601 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6661 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     6721 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     6781 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     6841 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     6901 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     6961 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     7021 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     7081 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     7141 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
