LOCUS       sequence-47452-e        7602 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 7602)
  AUTHORS   Birnbaum RY, Clowney EJ, Agamy O, Kim MJ, Zhao J, Yamanaka T, 
            Pappalardo Z, Clarke SL, Wenger AM, Nguyen L, Gurrieri F, Everman 
            DB, Schwartz CE, Birk OS, Bejerano G, Lomvardas S, Ahituv N
  TITLE     Coding exons function as tissue-specific enhancers of nearby genes.
  JOURNAL   Genome Res. 2012 Jun;22(6):1059-68. Epub 2012 Mar 22.
  PUBMED    22442009
REFERENCE   2  (bases 1 to 7602)
  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: "Genome 
            Res."; date: "2012-06"; volume: "22(6)"; pages: "1059-68. Epub 2012 
            Mar 22"
COMMENT     Sequence Label: E1b-GFP-Tol2-Gateway
FEATURES             Location/Qualifiers
     source          1..7602
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     49..68
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(287..391)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(417..872)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(554..575)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     766..785
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     999..1021
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1013..1030
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1014..1030
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1040..1059
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1040..1058
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(2908..2919,2920..3082)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     intron          3555..4127
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     regulatory      4201..4210
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4207..4209,4210..4212,4213..4926)
                     /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(4207..4209,4210..4212,4213..4923)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4252..4273)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4513..4532)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4860..4881
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4971..5105
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4995..5014)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5049..5068
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    join(5542..5729,5730..5741)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(5775..5791)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5775..5791)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(5828..5848)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5828..5846)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5867..5883)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5867..5883)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5880..5902)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5891..5907
                     /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(5915..5921,5922..5939,5940..5945))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5960..5981
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6098..6115)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6269..6857)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6349..6368)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7028..7602,1..217,218..286))
                     /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"
ORIGIN
        1 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
       61 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
      121 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
      181 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
      241 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
      301 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
      361 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat
      421 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
      481 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
      541 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
      601 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
      661 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
      721 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
      781 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
      841 cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg ccattcaggc tgcgcaactg
      901 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
      961 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     1021 gacggccagt gagcgcgcgt aatacgactc actatagggc gaattgggta ccaaatagta
     1081 ggaattaccc acctgtacaa gtgctgaaaa cttggatgaa taagcccgtt tgccattttc
     1141 tactgctatt ttaaatcttt tctgtatttg tctgcatttg tctttaccct tgaataaatg
     1201 ttttagttgt tttttttcaa ttatgactgt gtttaacaga cattattaca ataatatgta
     1261 atagtagtac acactattat tatgtaatag tacttgttga ctgtatttga gaactggacc
     1321 gagtgagtgt tacgtcaccc attcaaaatg acttacttct ggctccaaca aaatgaagtt
     1381 aattcagttg ccatttttca ctgtatggac atcgccgtgt tggagctaga cgttgccaat
     1441 aagcaagaaa gagccgagat gcgtcgagtc tgagtcaccg ttcctatggc aacccctcta
     1501 accaatcaga agtaagcttg ttggaagtcc acagcctacc acttgaaagc gggctgcaca
     1561 aaatctgtca aacgttttga acgttggatg tgagagcaca tacttttatt aaggcatctg
     1621 gttggtcagt ttataatatc aacaacttgg gctacagaaa agaaaagtta ttacagaaat
     1681 tatgattaac aagtacatgt taaataaaga ttttaatatg aatgccacca ctggagcatt
     1741 catgccattt ggagcttctt cctgtttgga tcactagaag gaggaggtca ctcattacag
     1801 ttctcatata cagtcgttgg ttggttggtt ggttggtaga ttgattgatt gattgattga
     1861 ttgattgatt gattgattga ttgattgatt gattgattga ttgattgatt gattggtagt
     1921 caaaataaga aataatttcc acagattcat tacagaaatg attaaatgca tacataaaaa
     1981 actggggggg gggggataca acaacacact taagtcacat ttgcctacgt aaagaaaagt
     2041 aaagaaaatc aatagctata ttttacatct cctttttttg ctgtctttta attagccttg
     2101 ttttgctgtt atcttgattg aaactgtaac ttcttcacct gcttcttttc tttgtaggtt
     2161 ttgccagccc agaaacaggc atggctgtgc aaggccagag caccatgcac aatgcgctgc
     2221 atgtcttcat gaacggctca atgtcctcag tccagggctc agccaacgac cccatcttcc
     2281 tccttcacca tgctttcatt gacaggtaac aaacacgtca tgacattaga ctgcacagtt
     2341 tttgacaaag ttcatacaat ctgttgttta tagctgctac aattagtgaa gtttgtgaat
     2401 gtacttggat gagcagcgaa agatcaattg agatcaattg ttagagtttg gttgccctgc
     2461 agagcaaaga acaaaaaata atctggtggc tttactgcgt gaggttatta ttggtggaat
     2521 agaaacacaa aacataattg catttatttg tttaattttt tatcttatct taactttcat
     2581 cttgcatatt tgtttcttac atcatttcta gcatctttga gcgctggcta agaactcatc
     2641 agcctccccg gtccatctac ccacgtacca atgcaccaat tggccacaat gacggctact
     2701 acatggtgcc attccttcct ctttatagga atggagacta cctcctgtcc aacaatgctc
     2761 ttggatacga gtacgcctac ctgttggacc caggtcattg cacaacacca gaaatgccct
     2821 ctgatctgca aaagacgtga atatctgttc agacacccat atccactctg ttccacacag
     2881 gtcagaggtt tgtccaggag ttcttgacag aggtgtaaaa agtactcaaa aattttactc
     2941 aagtgaaagt acaagtactt agggaaaatt ttactcaatt aaaagtaaaa gtatctggct
     3001 agaatcttac ttgagtaaaa gtaaaaaagt actccattaa aattgtactt gagtattaag
     3061 gaagtaaaag taaaagcaag aaagaaaact agagattctt gtttaagctt ttaatctcaa
     3121 aaaacattaa atgaaatgca tacaaggttt tatcctgctt tagaactgtt tgtatttaat
     3181 tatcaaacta taagacagac aatctaatgc cagtacacgc tactcaaagt tgtaaaacct
     3241 cagatttaac ttcagtagaa gctgattctc aaaattgtta gtgtcaagcc tagctctttt
     3301 ggggctgaaa agcaatcctg cagtgctgaa aagcctctca caggcagccg atgcgggaag
     3361 aggtgtatta gtcttgatag agaggctgca aatagcagga aacgtgagca gagactccct
     3421 ggtgtctgaa acacaggcca gatgggccct cgagagatct ctcgactcta gagggtatat
     3481 aatggatccc atcgcgtctc agcctcactt tgagctcctc cacacgaatt tcgaccgatc
     3541 ctgagaactt cagggtgagt ttggggaccc ttgattgttc tttctttttc gctattgtaa
     3601 aattcatgtt atatggaggg ggcaaagttt tcagggtgtt gtttagaatg ggaagatgtc
     3661 ccttgtatca ccatggaccc tcatgataat tttgtttctt tcactttcta ctctgttgac
     3721 aaccattgtc tcctcttatt ttcttttcat tttctgtaac tttttcgtta aactttagct
     3781 tgcatttgta acgaattttt aaattcactt ttgtttattt gtcagattgt aagtactttc
     3841 tctaatcact tttttttcaa ggcaatcagg gtatattata ttgtacttca gcacagtttt
     3901 agagaacaat tgttataatt aaatgataag gtagaatatt tctgcatata aattctggct
     3961 ggcgtggaaa tattcttatt ggtagaaaca actacaccct ggtcatcatc ctgcctttct
     4021 ctttatggtt acaatgatat acactgtttg agatgaggat aaaatactct gagtccaaac
     4081 cgggcccctc tgctaaccat gttcatgcct tcttctcttt cctacagctc ctgggcaacg
     4141 tgctggttgt tgtgctgtct catcattttg gcaaagaatt cctcgacgga tccaccggtc
     4201 gccaccatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
     4261 ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc
     4321 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
     4381 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
     4441 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc
     4501 atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
     4561 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
     4621 gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag
     4681 aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag
     4741 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
     4801 aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac
     4861 atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac
     4921 aagtaaagcg gccgccaccg cggtggagct cgaattaatt catcgatgat gatccagaca
     4981 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     5041 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     5101 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg
     5161 ttttttaaag caagtaaaac ctctacaaat gtggtatggc tgattatgat cctctagatc
     5221 agatccgaag atacggccac gggtgctctt gatcctgtgg ctgattttgg actgtgctgc
     5281 tcgcagctgc tgatgaatca catacttcct ccattttctt ccactgattg actgttataa
     5341 tttccctaat ttccaggtca aggtgctgtg cattgtggta atagatgtga catgacgtca
     5401 cttccaaagg accaatgaac atgtctgacc aatttcatat aatgtgaaaa cgattttcat
     5461 aggcagaata aataacattt aaattaaact gggcatcagc gcaattcaat tggtttggta
     5521 atagcaaggg aaaatagaat gaagtgatct ccaaaaaata agtacttttt gactgtaaat
     5581 aaaattgtaa ggagtaaaaa gtactttttt ttctaaaaaa atgtaattaa gtaaaagtaa
     5641 aagtattgat ttttaattgt actcaagtaa agtaaaaatc cccaaaaata atacttaagt
     5701 acagtaatca agtaaaatta ctcaagtact ttacacctct ggttcttgac cccctacctt
     5761 cagcaagccc agcagatcca ctagttctag agcggccgcc accgcggtgg agctccagct
     5821 tttgttccct ttagtgaggg ttaattgcgc gcttggcgta atcatggtca tagctgtttc
     5881 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     5941 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     6001 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     6061 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     6121 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     6181 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     6241 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     6301 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     6361 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     6421 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     6481 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     6541 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     6601 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     6661 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     6721 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     6781 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     6841 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     6901 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     6961 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     7021 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     7081 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     7141 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     7201 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     7261 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     7321 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     7381 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     7441 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     7501 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     7561 gcttttctgt gactggtgag tactcaacca agtcattctg ag
//
