LOCUS       sequence-334556-        6145 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Unregulated ""open-loop"" UBC promoter plasmid. This plasmid encodes
            eGFP to report circuit output levels..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6145)
  AUTHORS   Yang J, Lee J, Land MA, Lai S, Igoshin OA, St-Pierre F
  TITLE     A synthetic circuit for buffering gene dosage variation between 
            individual mammalian cells.
  JOURNAL   Nat Commun. 2021 Jul 5;12(1):4132. doi: 10.1038/s41467-021-23889-0.
  PUBMED    34226556
REFERENCE   2  (bases 1 to 6145)
  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: "2021-07-5"; volume: "12(1):4132. doi"; pages: " 
            10.1038/s41467-021-23889-0"
COMMENT     Sequence Label: UBC plasmid
FEATURES             Location/Qualifiers
     source          1..6145
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        19..1230
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     1067..1086
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     regulatory      1278..1287
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1284..1286,1287..1289,1290..2003)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1284..1286,1287..1289,1290..2000)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1329..1350)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1590..1609)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1937..1958
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2097..2114)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2103..2327
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2460..2479)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2611..2658
                     /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             2666..3688
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2666..3688
                     /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    3818..3939
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3855..3874)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3909..3928
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3988..4004)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3988..4004)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4001..4023)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4012..4028
                     /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(4036..4042,4043..4060,4061..4066))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4081..4102
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4219..4236)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4390..4978)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4470..4489)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5149..5940,5941..6009))
                     /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     5772..5791
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6010..6114)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctcgg cctccgcgcc gggttttggc gcctcccgcg ggcgcccccc
       61 tcctcacggc gagcgctgcc acgtcagacg aagggcgcag cgagcgtcct gatccttccg
      121 cccggacgct caggacagcg gcccgctgct cataagactc ggccttagaa ccccagtatc
      181 agcagaagga cattttagga cgggacttgg gtgactctag ggcactggtt ttctttccag
      241 agagcggaac aggcgaggaa aagtagtccc ttctcggcga ttctgcggag ggatctccgt
      301 ggggcggtga acgccgatga ttatataagg acgcgccggg tgtggcacag ctagttccgt
      361 cgcagccggg atttgggtcg cggttcttgt ttgtggatcg ctgtgatcgt cacttggtga
      421 gtagcgggct gctgggctgg ccggggcttt cgtggccgcc gggccgctcg gtgggacgga
      481 agcgtgtgga gagaccgcca agggctgtag tctgggtccg cgagcaaggt tgccctgaac
      541 tgggggttgg ggggagcgca gcaaaatggc ggctgttccc gagtcttgaa tggaagacgc
      601 ttgtgaggcg ggctgtgagg tcgttgaaac aaggtggggg gcatggtggg cggcaagaac
      661 ccaaggtctt gaggccttcg ctaatgcggg aaagctctta ttcgggtgag atgggctggg
      721 gcaccatctg gggaccctga cgtgaagttt gtcactgact ggagaactcg gtttgtcgtc
      781 tgttgcgggg gcggcagtta tggcggtgcc gttgggcagt gcacccgtac ctttgggagc
      841 gcgcgccctc gtcgtgtcgt gacgtcaccc gttctgttgg cttataatgc agggtggggc
      901 cacctgccgg taggtgtgcg gtaggctttt ctccgtcgca ggacgcaggg ttcgggccta
      961 gggtaggctc tcctgaatcg acaggcgccg gacctctggt gaggggaggg ataagtgagg
     1021 cgtcagtttc tttggtcggt tttatgtacc tatcttctta agtagctgaa gctccggttt
     1081 tgaactatgc gctcggggtt ggcgagtgtg ttttgtgaag ttttttaggc accttttgaa
     1141 atgtaatcat ttgggtcaat atgtaatttt cagtgttaga ctagtaaatt gtccgctaaa
     1201 ttctggccgt ttttggcttt tttgttagac gaagcttggg ctgcaggtcg actctagagg
     1261 atccccgggt accggtcgcc accatggtga gcaagggcga ggagctgttc accggggtgg
     1321 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     1381 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
     1441 agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca
     1501 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     1561 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     1621 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     1681 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     1741 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg
     1801 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     1861 ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca
     1921 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     1981 gcatggacga gctgtacaag tagggatcca ctagtccagt gtggtggaat tctgcagata
     2041 tccagcacag tggcggccgc tcgagtctag agggcccgtt taaacccgct gatcagcctc
     2101 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     2161 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     2221 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     2281 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     2341 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     2401 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     2461 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     2521 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     2581 acttgattag ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct
     2641 agaaagtata ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa
     2701 gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga
     2761 atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg
     2821 cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc
     2881 gattccggaa gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg
     2941 ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca
     3001 gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt
     3061 cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc
     3121 gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc
     3181 cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca
     3241 cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc
     3301 ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt
     3361 cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca
     3421 tccggagctt gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca
     3481 actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg
     3541 cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag
     3601 cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc
     3661 cagcactcgt ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc
     3721 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     3781 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
     3841 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     3901 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
     3961 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     4021 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     4081 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     4141 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     4201 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     4261 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     4321 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     4381 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     4441 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     4501 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     4561 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     4621 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     4681 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     4741 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     4801 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     4861 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     4921 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4981 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     5041 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     5101 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     5161 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     5221 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     5281 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     5341 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     5401 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     5461 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     5521 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     5581 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     5641 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     5701 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     5761 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     5821 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     5881 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     5941 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     6001 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     6061 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     6121 tccccgaaaa gtgccacctg acgtc
//
