LOCUS       sequence-61270-e        6823 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6823)
  AUTHORS   Nevozhay D, Zal T, Balazsi G
  TITLE     Transferring a synthetic gene circuit from yeast to mammalian cells.
  JOURNAL   Nat Commun. 2013 Feb 5;4:1451. doi: 10.1038/ncomms2471.
  PUBMED    23385595
REFERENCE   2  (bases 1 to 6823)
  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: "2013-02-5"; volume: "4:1451. doi"; pages: " 
            10.1038/ncomms2471"
COMMENT     Sequence Label: pDN-D2irC6kwh
FEATURES             Location/Qualifiers
     source          1..6823
                     /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    814..832
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    840..858
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     intron          1041..1613
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     regulatory      1625..1634
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1631..2341
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1776..1794)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2044..2063
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2409..2429)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     primer_bind     complement(2782..2799)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2788..3012
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3058..3486
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3145..3164)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3355..3376
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3495..3515)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3500..3829
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3680..3815
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3742..3761
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3877..3924
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3943..4341
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4499..4620
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4536..4555)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4590..4609
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4669..4685)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4669..4685)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4682..4704)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4693..4709
                     /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(4717..4723,4724..4741,4742..4747))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4762..4783
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4900..4917)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5071..5656)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5151..5170)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5827..6618,6619..6687))
                     /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     6450..6469
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6688..6792)
                     /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 gcatccctat cagtgataga gatcagatct
      841 ccctatcagt gatagagagc tgttttgacc tccatagaag acaccgggac cgatccagcc
      901 tccggactct agcgtttaaa cttaagcttg gtacccgggg atcctctagg gcctctgagc
      961 tattccagaa gtagtgaaga ggcttttttg gaggcctagg cttttgcaaa aagctccgga
     1021 tcgatcctga gaacttcagg gtgagtttgg ggacccttga ttgttctttc tttttcgcta
     1081 ttgtaaaatt catgttatat ggagggggca aagttttcag ggtgttgttt agaatgggaa
     1141 gatgtccctt gtatcaccat ggaccctcat gataattttg tttctttcac tttctactct
     1201 gttgacaacc attgtctcct cttattttct tttcattttc tgtaactttt tcgttaaact
     1261 ttagcttgca tttgtaacga atttttaaat tcacttttgt ttatttgtca gattgtaagt
     1321 actttctcta atcacttttt tttcaaggca atcagggtat attatattgt acttcagcac
     1381 agttttagag aacaattgtt ataattaaat gataaggtag aatatttctg catataaatt
     1441 ctggctggcg tggaaatatt cttattggta gaaacaacta catcctggtc atcatcctgc
     1501 ctttctcttt atggttacaa tgatatacac tgtttgagat gaggataaaa tactctgagt
     1561 ccaaaccggg cccctctgct aaccatgttc atgccttctt ctttttccta caggtcctgc
     1621 aggcgccacc atggtgagca agggcgagga ggataacatg gccatcatca aggagttcat
     1681 gcgcttcaag gtgcacatgg agggctccgt gaacggccac gagttcgaga tcgagggcga
     1741 gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg
     1801 ccccctgccc ttcgcctggg acatcctgtc ccctcagttc atgtacggct ccaaggccta
     1861 cgtgaagcac cccgccgaca tccccgacta cttgaagctg tccttccccg agggcttcaa
     1921 gtgggagcgc gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc
     1981 cctgcaggac ggcgagttca tctacaaggt gaagctgcgc ggcaccaact tcccctccga
     2041 cggccccgta atgcagaaga agaccatggg ctgggaggcc tcctccgagc ggatgtaccc
     2101 cgaggacggc gccctgaagg gcgagatcaa gcagaggctg aagctgaagg acggcggcca
     2161 ctacgacgct gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc
     2221 ctacaacgtc aacatcaagt tggacatcac ctcccacaac gaggactaca ccatcgtgga
     2281 acagtacgaa cgcgccgagg gccgccactc caccggcggc atggacgagc tgtacaagta
     2341 gaccggtgcg gccgcaatca acctctggat tacaaaattt gtgaaagatt gactggtatt
     2401 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
     2461 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
     2521 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
     2581 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
     2641 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
     2701 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcctcgag tctagagggc
     2761 ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     2821 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     2881 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     2941 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     3001 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
     3061 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     3121 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     3181 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     3241 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     3301 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     3361 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     3421 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     3481 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     3541 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     3601 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     3661 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     3721 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     3781 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     3841 agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag
     3901 tatatcggca tagtataata cgacaaggtg aggaactaaa ccatggccaa gcctttgtct
     3961 caagaagaat ccaccctcat tgaaagagca acggctacaa tcaacagcat ccccatctct
     4021 gaagactaca gcgtcgccag cgcagctctc tctagcgacg gccgcatctt cactggtgtc
     4081 aatgtatatc attttactgg gggaccttgt gcagaactcg tggtgctggg cactgctgct
     4141 gctgcggcag ctggcaacct gacttgtatc gtcgcgatcg gaaatgagaa caggggcatc
     4201 ttgagcccct gcggacggtg ccgacaggtg cttctcgatc tgcatcctgg gatcaaagcc
     4261 atagtgaagg acagtgatgg acagccgacg gcagttggga ttcgtgaatt gctgccctct
     4321 ggttatgtgt gggagggcta agcacttcgt ggccgaggag caggactgac acgtgctacg
     4381 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     4441 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     4501 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     4561 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     4621 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     4681 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     4741 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     4801 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     4861 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     4921 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     4981 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5041 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5101 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5161 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5221 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     5281 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5341 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5401 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     5461 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     5521 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     5581 cggcaaacaa accaccgctg gtagcggttt ttttgtttgc aagcagcaga ttacgcgcag
     5641 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5701 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5761 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5821 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5881 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5941 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6001 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6061 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6121 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6181 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6241 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6301 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6361 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6421 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6481 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6541 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6601 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6661 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6721 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6781 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
