LOCUS       sequence-233156-        7827 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ePiggyBac mammalian expression plasmid for TetR-tdTom fusion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7827)
  AUTHORS   Alexander JM, Guan J, Li B, Maliskova L, Song M, Shen Y, Huang B, 
            Lomvardas S, Weiner OD
  TITLE     Live-cell imaging reveals enhancer-dependent Sox2 transcription in 
            the absence of enhancer proximity.
  JOURNAL   Elife. 2019 May 24;8. pii: 41769. doi: 10.7554/eLife.41769.
  PUBMED    31124784
REFERENCE   2  (bases 1 to 7827)
  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: "Elife."; 
            date: "2019-05-24"; volume: "8. pii: 41769. doi"; pages: " 
            10.7554/eLife.41769"
COMMENT     Sequence Label: epB_CAG_TetRFlag-nls-tdTom_DEx4
FEATURES             Location/Qualifiers
     source          1..7827
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        137..516
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          796..1812
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1736..1758
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1820..1839
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    1879..1903
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1965..2588
                     /codon_start=1
                     /gene="tetR"
                     /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="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(2041..2060)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     CDS             2589..2612
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2691..4121
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     protein_bind    complement(4153..4177)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    4360..4481
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4397..4416)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4451..4470
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   4667..4729
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(4764..4783)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4765..4783)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4793..4810)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4793..4809)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4802..4824)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4951..5406
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5038..5057)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5248..5269
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5432..5536
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5537..5605,5606..6397)
                     /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     complement(5755..5774)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6568..7156
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7057..7076
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7310..7327
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7444..7465
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7480..7485,7486..7503,7504..7510)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7518..7534
                     /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)."
     primer_bind     7523..7545
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7542..7558
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7542..7558
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7577..7597
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7579..7597
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     repeat_region   7645..7679
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
ORIGIN
        1 tcaaaatgac gcatgattat cttttacgtg acttttaaga tttaactcat acgataatta
       61 tattgttatt tcgtgttcta cttacgtgat aacttattat atatatattt tcttgttata
      121 gatatccttc tcgagcgaca ttgattattg actagttatt aatagtaatc aattacgggg
      181 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
      241 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
      301 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
      361 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
      421 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
      481 cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc cacgttctgc
      541 ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt tattttttaa
      601 ttattttgtg cagcgatggg ggcggggggg gggggggggc gcgcgccrgg sggggsgggg
      661 sggggsgrgg ggsggggsgg ggsgrggsgg rgaggtgcgg cggcagccaa tcagagcggc
      721 gcgctccgaa agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga
      781 agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg ctccgccgcc
      841 gcctcgcgcc gcccgccccg gctctgactg accgcgttac tcccacaggt gagcgggcgg
      901 gacggccctt ctcctccggg ctgtaattag cgcttggttt aatgacggct tgtttctttt
      961 ctgtggctgc gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg ggggagcggc
     1021 tcggggggtg cgtgcgtgtg tgtgtgcgtg gggagcgccg cgtgcggctc cgcgctgccc
     1081 ggcggctgtg agcgctgcgg gcgcggcgcg gggctttgtg cgctccgcag tgtgcgcgag
     1141 gggagcgcgg ccgggggcgg tgccccgcgg tgcggggggg gctgcgaggg gaacaaaggc
     1201 tgcgtgcggg gtgtgtgcgt gggggggtga gcagggggtg tgggcgcgtc ggtcgggctg
     1261 caaccccccc tgcacccccc tccccgagtt gctgagcacg gcccggcttc gggtgcgggg
     1321 ctccgtacgg ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg gcaggtgggg
     1381 gtgccgggcg gggcggggcc gcctcgggcc ggggagggct cgggggaggg gcgcggcggc
     1441 ccccggagcg ccggcggctg tcgaggcgcg gcgagccgca gccattgcct tttatggtaa
     1501 tcgtgcgaga gggcgcaggg acttcctttg tcccaaatct gtgcggagcc gaaatctggg
     1561 aggcgccgcc gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag
     1621 gaaatgggcg gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag
     1681 cctcggggct gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc
     1741 ggcttctggc gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc
     1801 tttttcctac agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa
     1861 gaattgctcg aggatatcac aagtttgtac aaaaaagcag gctttaaagg aaccaattca
     1921 gtcgactgga tccggtaccg aattcgcggc cgcctagcta aaccatgatg tctagattag
     1981 ataaaagtaa agtgattaac agcgcattag agctgcttaa tgaggtcgga atcgaaggtt
     2041 taacaacccg taaactcgcc cagaagctag gtgtagagca gcctacattg tattggcatg
     2101 taaaaaataa gcgggctttg ctcgacgcct tagccattga gatgttagat aggcaccata
     2161 ctcacttttg ccctttagaa ggggaaagct ggcaagattt tttacgtaat aacgctaaaa
     2221 gttttagatg tgctttacta agtcatcgcg atggagcaaa agtacattta ggtacacggc
     2281 ctacagaaaa acagtatgaa actctcgaaa atcaattagc ctttttatgc caacaaggtt
     2341 tttcactaga gaatgcatta tatgcactca gcgctgtggg gcattttact ttaggttgcg
     2401 tattggaaga tcaagagcat caagtcgcta aagaagaaag ggaaacacct actactgata
     2461 gtatgccgcc attattacga caagctatcg aattatttga tcaccaaggt gcagagccag
     2521 ccttcttatt cggccttgaa ttgatcatat gcggattaga aaaacaactt aaatgtgaaa
     2581 gtgggtctga ttataaagat gatgatgata aaccggataa ggcacgtcgt ccagcagtaa
     2641 gatcaccacc actaggaact cgtaagcgaa agcgtccttc tcggggtctg atggtgagca
     2701 agggcgagga ggtcatcaaa gagttcatgc gcttcaaggt gcgcatggag ggctccatga
     2761 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
     2821 ccgccaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac atcctgtccc
     2881 cccagttcat gtacggctcc aaggcgtacg tgaagcaccc cgccgacatc cccgattaca
     2941 agaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     3001 gtctggtgac cgtgacccag gactcctccc tgcaggacgg cacgctgatc tacaaggtga
     3061 agatgcgcgg caccaacttc ccccccgacg gccccgtaat gcagaagaag accatgggct
     3121 gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc gagatccacc
     3181 aggccctgaa gctgaaggac ggcggccact acctggtgga gttcaagacc atctacatgg
     3241 ccaagaagcc cgtgcaactg cccggctact actacgtgga caccaagctg gacatcacct
     3301 cccacaacga ggactacacc atcgtggaac agtacgagcg ctccgagggc cgccaccacc
     3361 tgttcctggg gcatggcacc ggcagcaccg gcagcggcag ctccggcacc gcctcctccg
     3421 aggacaacaa catggccgtc atcaaagagt tcatgcgctt caaggtgcgc atggagggct
     3481 ccatgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgagggca
     3541 cccagaccgc caagctgaag gtgaccaagg gcggccccct gcccttcgcc tgggacatcc
     3601 tgtcccccca gttcatgtac ggctccaagg cgtacgtgaa gcaccccgcc gacatccccg
     3661 attacaagaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg aacttcgagg
     3721 acggcggtct ggtgaccgtg acccaggact cctccctgca ggacggcacg ctgatctaca
     3781 aggtgaagat gcgcggcacc aacttccccc ccgacggccc cgtaatgcag aagaagacca
     3841 tgggctggga ggcctccacc gagcgcctgt acccccgcga cggcgtgctg aagggcgaga
     3901 tccaccaggc cctgaagctg aaggacggcg gccactacct ggtggagttc aagaccatct
     3961 acatggccaa gaagcccgtg caactgcccg gctactacta cgtggacacc aagctggaca
     4021 tcacctccca caacgaggac tacaccatcg tggaacagta cgagcgctcc gagggccgcc
     4081 accacctgtt cctgtacggc atggacgagc tgtacaagta agtgcctagg cggccgcact
     4141 cgagatatct agacccagct ttcttgtaca aagtggtgat ccttatttat tgatccttat
     4201 ttattgtagc ttatttattg ctacttattt attgatctgc ggccgcgact ctagatcata
     4261 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     4321 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     4381 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     4441 cattctagtt gtggtttgtc caaactcatc aatgtatctt aaggaattcg ataaaagttt
     4501 tgttacttta tagaagaaat tttgagtttt tgtttttttt taataaataa ataaacataa
     4561 ataaattgtt tgttgaattt attattagta tgtaagtgta aatataataa aacttaatat
     4621 ctattcaaat taataaataa acctcgatat acagaccgat aaaacacatg cgtcaatttt
     4681 acgcatgatt atctttaacg tacgtcacaa tatgattatc tttctagggt taatctagct
     4741 gcggcgcgcc ggtacccaat tcgccctata gtgagtcgta ttacgcgcgc tcactggccg
     4801 tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag
     4861 cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
     4921 aacagttgcg cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
     4981 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     5041 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     5101 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     5161 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     5221 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     5281 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     5341 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     5401 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     5461 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     5521 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     5581 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     5641 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     5701 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     5761 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     5821 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     5881 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     5941 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     6001 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     6061 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     6121 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     6181 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     6241 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     6301 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     6361 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     6421 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     6481 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     6541 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     6601 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     6661 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     6721 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     6781 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     6841 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     6901 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     6961 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     7021 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     7081 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     7141 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     7201 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     7261 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     7321 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     7381 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
     7441 aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc
     7501 tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca
     7561 tgattacgcc aagcgcgcaa ttaaccctca ctaaagggaa caaaagctgg agctccaccg
     7621 cggggcgcgc cgcagctaga ttaaccctag aaagatagtc tgcgtaaaat tgacgcatgc
     7681 attcttgaaa tattgctctc tctttctaaa tagcgcgaat ccgtcgctgt gcatttagga
     7741 catctcagtc gccgcttgga gctcccgtga ggcgtgcttg tcaatgcggt aagtgtcact
     7801 gattttgaac tataacaacc gcgtgag
//
