LOCUS       sequence-232743-        7697 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ePiggyBac mammalian expression plasmid for CymR-Halo fusion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7697)
  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 7697)
  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_UbC_CymRV5-nls-Halox2_DEx4
FEATURES             Location/Qualifiers
     source          1..7697
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        98..1309
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     1146..1165
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     protein_bind    1326..1350
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1412..2020
                     /codon_start=1
                     /gene="cymR from Pseudomonas putida"
                     /product="cumate repressor (Mullick et al., 2006)"
                     /label=CymR
                     /translation="MSPKRRTQAERAMETQGKLIAAALGVLREKGYAGFRIADVPGAAG
                     VSRGAQSHHFPTKLELLLATFEWLYEQITERSRARLAKLKPEDDVIQQMLDDAAEFFLD
                     DDFSISLDLIVAADRDPALREGIQRTVERNRFVVEDMWLGVLVSRGLSRDDAEDILWLI
                     FNSVRGLAVRSLWQKDKERFERVRNSTLEIARERYAKFKR"
     CDS             2021..2062
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2141..3031
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             3038..3928
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     protein_bind    complement(3963..3987)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    4170..4291
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4207..4226)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4261..4280
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   4477..4539
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(4574..4593)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4575..4593)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4603..4620)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4603..4619)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4612..4634)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4761..5216
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4848..4867)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5058..5079
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5242..5346
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5347..5415,5416..6207)
                     /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(5565..5584)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6378..6966
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6867..6886
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7120..7137
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7254..7275
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7290..7295,7296..7313,7314..7320)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7328..7344
                     /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     7333..7355
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7352..7368
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7352..7368
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7387..7407
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7389..7407
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     repeat_region   7455..7489
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
ORIGIN
        1 tattgttatt tcgtgttcta cttacgtgat aacttattat atatatattt tcttgttata
       61 gatatccttc tcgacattga ttattgacta gagatctggc ctccgcgccg ggttttggcg
      121 cctcccgcgg gcgcccccct cctcacggcg agcgctgcca cgtcagacga agggcgcagc
      181 gagcgtcctg atccttccgc ccggacgctc aggacagcgg cccgctgctc ataagactcg
      241 gccttagaac cccagtatca gcagaaggac attttaggac gggacttggg tgactctagg
      301 gcactggttt tctttccaga gagcggaaca ggcgaggaaa agtagtccct tctcggcgat
      361 tctgcggagg gatctccgtg gggcggtgaa cgccgatgat tatataagga cgcgccgggt
      421 gtggcacagc tagttccgtc gcagccggga tttgggtcgc ggttcttgtt tgtggatcgc
      481 tgtgatcgtc acttggtgag tagcgggctg ctgggctggc cggggctttc gtggccgccg
      541 ggccgctcgg tgggacggaa gcgtgtggag agaccgccaa gggctgtagt ctgggtccgc
      601 gagcaaggtt gccctgaact gggggttggg gggagcgcag caaaatggcg gctgttcccg
      661 agtcttgaat ggaagacgct tgtgaggcgg gctgtgaggt cgttgaaaca aggtgggggg
      721 catggtgggc ggcaagaacc caaggtcttg aggccttcgc taatgcggga aagctcttat
      781 tcgggtgaga tgggctgggg caccatctgg ggaccctgac gtgaagtttg tcactgactg
      841 gagaactcgg tttgtcgtct gttgcggggg cggcagttat ggcggtgccg ttgggcagtg
      901 cacccgtacc tttgggagcg cgcgccctcg tcgtgtcgtg acgtcacccg ttctgttggc
      961 ttataatgca gggtggggcc acctgccggt aggtgtgcgg taggcttttc tccgtcgcag
     1021 gacgcagggt tcgggcctag ggtaggctct cctgaatcga caggcgccgg acctctggtg
     1081 aggggaggga taagtgaggc gtcagtttct ttggtcggtt ttatgtacct atcttcttaa
     1141 gtagctgaag ctccggtttt gaactatgcg ctcggggttg gcgagtgtgt tttgtgaagt
     1201 tttttaggca ccttttgaaa tgtaatcatt tgggtcaata tgtaattttc agtgttagac
     1261 tagtaaattg tccgctaaat tctggccgtt tttggctttt ttgttagacg aagctcgagg
     1321 atatcacaag tttgtacaaa aaagcaggct ttaaaggaac caattcagtc gactggatcc
     1381 ggtaccgaat tcgcggccgc ctagctaaac catgtcacct aagagaagaa ctcaggcaga
     1441 gagagcaatg gaaactcagg ggaaactcat cgcagccgca ctcggggtcc tcagagaaaa
     1501 gggctacgca ggattccgga tcgccgatgt gccaggagca gctggtgtct cacgcggggc
     1561 tcagagccac catttcccta ccaaactgga gctgctcctg gcaacatttg agtggctgta
     1621 tgaacagatc actgagagga gcagagcccg gctcgctaag ctgaaaccag aggacgatgt
     1681 gattcagcag atgctcgacg atgcagccga attctttctg gacgatgact tcagcatctc
     1741 cctcgatctg attgtggctg cagatcggga ccccgccctg cgagaaggga ttcagcgaac
     1801 cgtggagcgc aaccgatttg tggtcgaaga tatgtggctc ggcgtgctgg tgtctagggg
     1861 actgagtaga gatgacgccg aggacatcct ctggctgatt ttcaattccg tgaggggcct
     1921 cgctgtcaga tctctgtggc agaaggacaa agagcggttt gaacgagtca gaaacagcac
     1981 actggaaatc gcacgggaga gatacgctaa gttcaaaagg ggtaagccta tccctaaccc
     2041 tctcctcggt ctcgattcta cgccggataa ggcacgtcgt ccagcagtaa gatcaccacc
     2101 actaggaact cgtaagcgaa agcgtccttc tcggggtctg atggcagaaa tcggtactgg
     2161 ctttccattc gacccccatt atgtggaagt cctgggcgag cgcatgcact acgtcgatgt
     2221 tggtccgcgc gatggcaccc ctgtgctgtt cctgcacggt aacccgacct cctcctacgt
     2281 gtggcgcaac atcatcccgc atgttgcacc gacccatcgc tgcattgctc cagacctgat
     2341 cggtatgggc aaatccgaca aaccagacct gggttatttc ttcgacgacc acgtccgctt
     2401 catggatgcc ttcatcgaag ccctgggtct ggaagaggtc gtcctggtca ttcacgactg
     2461 gggctccgct ctgggtttcc actgggccaa gcgcaatcca gagcgcgtca aaggtattgc
     2521 atttatggag ttcatccgcc ctatcccgac ctgggacgaa tggccagaat ttgcccgcga
     2581 gaccttccag gccttccgca ccaccgacgt cggccgcaag ctgatcatcg atcagaacgt
     2641 ttttatcgag ggtacgctgc cgatgggtgt cgtccgcccg ctgactgaag tcgagatgga
     2701 ccattaccgc gagccgttcc tgaatcctgt tgaccgcgag ccactgtggc gcttcccaaa
     2761 cgagctgcca atcgccggtg agccagcgaa catcgtcgcg ctggtcgaag aatacatgga
     2821 ctggctgcac cagtcccctg tcccgaagct gctgttctgg ggcaccccag gcgttctgat
     2881 cccaccggcc gaagccgctc gcctggccaa aagcctgcct aactgcaagg ctgtggacat
     2941 cggcccgggt ctgaatctgc tgcaagaaga caacccggac ctgatcggca gcgagatcgc
     3001 gcgctggctg tctactctgg agatttccgg cggatctatg gcagaaatcg gtactggctt
     3061 tccattcgac ccccattatg tggaagtcct gggcgagcgc atgcactacg tcgatgttgg
     3121 tccgcgcgat ggcacccctg tgctgttcct gcacggtaac ccgacctcct cctacgtgtg
     3181 gcgcaacatc atcccgcatg ttgcaccgac ccatcgctgc attgctccag acctgatcgg
     3241 tatgggcaaa tccgacaaac cagacctggg ttatttcttc gacgaccacg tccgcttcat
     3301 ggatgccttc atcgaagccc tgggtctgga agaggtcgtc ctggtcattc acgactgggg
     3361 ctccgctctg ggtttccact gggccaagcg caatccagag cgcgtcaaag gtattgcatt
     3421 tatggagttc atccgcccta tcccgacctg ggacgaatgg ccagaatttg cccgcgagac
     3481 cttccaggcc ttccgcacca ccgacgtcgg ccgcaagctg atcatcgatc agaacgtttt
     3541 tatcgagggt acgctgccga tgggtgtcgt ccgcccgctg actgaagtcg agatggacca
     3601 ttaccgcgag ccgttcctga atcctgttga ccgcgagcca ctgtggcgct tcccaaacga
     3661 gctgccaatc gccggtgagc cagcgaacat cgtcgcgctg gtcgaagaat acatggactg
     3721 gctgcaccag tcccctgtcc cgaagctgct gttctggggc accccaggcg ttctgatccc
     3781 accggccgaa gccgctcgcc tggccaaaag cctgcctaac tgcaaggctg tggacatcgg
     3841 cccgggtctg aatctgctgc aagaagacaa cccggacctg atcggcagcg agatcgcgcg
     3901 ctggctgtct actctggaga tttccggcta agtgcctagg cggccgcact cgagatatct
     3961 agacccagct ttcttgtaca aagtggtgat ccttatttat tgatccttat ttattgtagc
     4021 ttatttattg ctacttattt attgatctgc ggccgcgact ctagatcata atcagccata
     4081 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
     4141 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
     4201 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     4261 gtggtttgtc caaactcatc aatgtatctt aaggaattcg ataaaagttt tgttacttta
     4321 tagaagaaat tttgagtttt tgtttttttt taataaataa ataaacataa ataaattgtt
     4381 tgttgaattt attattagta tgtaagtgta aatataataa aacttaatat ctattcaaat
     4441 taataaataa acctcgatat acagaccgat aaaacacatg cgtcaatttt acgcatgatt
     4501 atctttaacg tacgtcacaa tatgattatc tttctagggt taatctagct gcggcgcgcc
     4561 ggtacccaat tcgccctata gtgagtcgta ttacgcgcgc tcactggccg tcgttttaca
     4621 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     4681 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     4741 cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     4801 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     4861 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     4921 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     4981 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     5041 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     5101 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     5161 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt
     5221 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     5281 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     5341 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     5401 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     5461 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     5521 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     5581 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     5641 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     5701 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     5761 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     5821 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     5881 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     5941 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     6001 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     6061 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     6121 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     6181 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     6241 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     6301 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     6361 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     6421 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     6481 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     6541 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     6601 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     6661 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     6721 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     6781 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     6841 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     6901 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     6961 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     7021 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     7081 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     7141 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     7201 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg
     7261 agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg
     7321 tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc
     7381 aagcgcgcaa ttaaccctca ctaaagggaa caaaagctgg agctccaccg cggggcgcgc
     7441 cgcagctaga ttaaccctag aaagatagtc tgcgtaaaat tgacgcatgc attcttgaaa
     7501 tattgctctc tctttctaaa tagcgcgaat ccgtcgctgt gcatttagga catctcagtc
     7561 gccgcttgga gctcccgtga ggcgtgcttg tcaatgcggt aagtgtcact gattttgaac
     7621 tataacaacc gcgtgagtca aaatgacgca tgattatctt ttacgtgact tttaagattt
     7681 aactcatacg ataatta
//
