LOCUS       sequence-136084-        7438 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tol2 vector with Notch responsive tp1 element driving expression of 
            EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7438)
  AUTHORS   Quillien A, Moore JC, Shin M, Siekmann AF, Smith T, Pan L, Moens CB,
            Parsons MJ, Lawson ND
  TITLE     Distinct Notch signaling outputs pattern the developing arterial 
            system.
  JOURNAL   Development. 2014 Apr;141(7):1544-52. doi: 10.1242/dev.099986. Epub 
            2014 Mar 5.
  PUBMED    24598161
REFERENCE   2  (bases 1 to 7438)
  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"; doi: 
            "10.1242/dev.099986"; journalName: "Development"; date: "2014-04"; 
            volume: "141"; issue: "7"; pages: "1544-52"
COMMENT     Sequence Label: 625 pTol2-tp1bglob:egfp
FEATURES             Location/Qualifiers
     source          1..7438
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(34..462)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(144..165)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     356..375
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     589..611
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     603..620
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     604..620
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        630..648
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(778..789,790..952)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     protein_bind    1779..1803
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      1815..1824
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1821..1823,1824..1826,1827..2537)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1821..1823,1824..1826,1827..2537)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1866..1887)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2127..2146)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2474..2495
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2538..2603
                     /label=MCS
                     /note="multiple cloning site"
     protein_bind    complement(2624..2648)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    2693..2827
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2717..2736)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2771..2790
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    join(4968..5155,5156..5167)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(5201..5217)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5201..5217)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(5254..5274)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5254..5272)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5293..5309)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5293..5309)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5306..5328)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5317..5333
                     /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(5341..5347,5348..5365,5366..5371))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5386..5407
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5524..5541)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5695..6283)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5775..5794)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6454..7245,7246..7314))
                     /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     7077..7096
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7315..7419)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
       61 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
      121 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
      181 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
      241 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
      301 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
      361 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
      421 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg ccattcaggc
      481 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
      541 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
      601 gttgtaaaac gacggccagt gagcgcgcgt aatacgactc actatagggc gaattgggta
      661 caggtcattg cacaacacca gaaatgccct ctgatctgca aaagacgtga atatctgttc
      721 agacacccat atccactctg ttccacacag gtcagaggtt tgtccaggag ttcttgacag
      781 aggtgtaaaa agtactcaaa aattttactc aagtgaaagt acaagtactt agggaaaatt
      841 ttactcaatt aaaagtaaaa gtatctggct agaatcttac ttgagtaaaa gtaaaaaagt
      901 actccattaa aattgtactt gagtattaag gaagtaaaag taaaagcaag aaagaaaact
      961 agagattctt gtttaagctt ttaatctcaa aaaacattaa atgaaatgca tacaaggttt
     1021 tatcctgctt tagaactgtt tgtatttaat tatcaaacta taagacagac aatctaatgc
     1081 cagtacacgc tactcaaagt tgtaaaacct cagatttaac ttcagtagaa gctgattctc
     1141 aaaattgtta gtgtcaagcc tagctctttt ggggctgaaa agcaatcctg cagtgctgaa
     1201 aagcctctca caggcagccg atgcgggaag aggtgtatta gtcttgatag agaggctgca
     1261 aatagcagga aacgtgagca gagactccct ggtgtctgaa acacaggcca gatgggccga
     1321 tagcttgcat gcctgcaggt cgactctaga ggatcccgac tcatgggaaa atgggcggaa
     1381 gggcaccgtg ggaaaatagt agatcccgac tcgtgggaaa atgggcggaa gggcaccgtg
     1441 ggaaaatagt agatcccgac tcgtgggaaa atgggcggaa gggcaccgtg ggaaaatagt
     1501 agatcccgac tcatgggaaa atgggcggaa gggcaccgtg ggaaaatagt agatcccgac
     1561 tcgtgggaaa atgggcggaa gggcaccgtg ggaaaatagt agatcccgac tcgtgggaaa
     1621 atgggcggaa gggcaccgtg ggaaaatagt agatccactg tatgcatctt gggcataaaa
     1681 ggcagatcac ttcagcttct gcttacactt gcttttgaca caactgtgtt tacttgcaat
     1741 ccctcgaatc gctagcatcg attcgaattc aaggatcaac aagtttgtac aaaaaagcag
     1801 gctcgctacc ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
     1861 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     1921 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     1981 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
     2041 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     2101 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     2161 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     2221 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     2281 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     2341 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     2401 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     2461 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     2521 tggacgagct gtacaagtcc ggactcagat ctcgagctca agcttcgaat tctgcagtcg
     2581 acggtaccgc gggcccggga tccaccggat ctagataact gatacccagc tttcttgtac
     2641 aaagtggttg atcctctcga gcctctagaa ctatagtgag tcgtattacg tagatccaga
     2701 catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg
     2761 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     2821 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga
     2881 ggttttttaa ttcgcggccg cggcgccaat gcattgggcc ctcgagcata tgagatctgt
     2941 cccgatcttt ctcttctgtg ctgtcaattt agagtagttt ttgaggtaat ttgggtgcat
     3001 tctctgtaaa aacaacaaat aaatatttat attgtatgca ataaatgtaa caattacaca
     3061 tacactctct gattacagca ttaaagggtt agttcaccca aaaatgaaaa gatgtcatta
     3121 atgactcacc ctcatgtcgt tccaagcccg taacacctcc gttcatcttc ggaacacagt
     3181 ttaagatatt ttagatttag tccgagagct ttctgtccct ccactgaaaa tgtatgtacg
     3241 gtatactgtc catgtccaga aaggtaataa aaacatcaaa gtagtccatg tgacatcagt
     3301 gggttaggta gaattatttg aagcatcgaa aatacatttt ggtccaaaaa taacaaaacc
     3361 tacgacttta ttcagcattg tagcgtcact gcgaagtcgt gaacgcggat tgacaacaga
     3421 cccggaagag aatacaatgc cgaataaagt cgtaggtttt gttatttttg gaccaaaatg
     3481 tattcgatgc ttcaaaaaat tctaactaac ccactgatgt cacatggact actttgatgt
     3541 ttttattacc tttctggaca tggacagtat accgtacata cattctcaat ggaggcacag
     3601 aaagctctcg gactaaatct aaaatatctt aaactgtgtt ctgaagatga acggaggtct
     3661 tacgggcttg gaacgacatg agggcgagtc attaatgaca ttattttcat ttttgggtga
     3721 actaaccctt taataaaact gacatttgca gccaattgca cactttataa tgttaaatac
     3781 ttgttgattt tagaaataca gtagcattta gtgacaatca aacattaatt ttatttctga
     3841 ttatattgac aaaaacacag gactgtaagg ctgacatgac agggtgaatt gatttcatta
     3901 cattagtttt gatggtaaca cttttacaat aaggttcgtt agttaactac attagttaac
     3961 atgaactaat aatgaactgc acgtatacat gttaatttca acatttacta atactttata
     4021 tcttgttaac attagttaat gcactgtgaa ctaacatgaa caaacaatga acatctgtat
     4081 ttctattaac taacgttaac aaagattaac agatacagta acaaatgtat tgctcatggt
     4141 tatttaatgt tagttaatac atgaactaat caacctcatt gtaaagtgtt accgttttga
     4201 caatacaaca tgtttggaaa aaaatttctc gcttggtttt cttgtgtttt gcatgaattc
     4261 aaacacaatg agagcacgga atgataatga ttattcaaag ctgcaatagg cgccaaattt
     4321 cccgtggacc taacgttacc aattacaaat gcattttagc attttattac gtgagcctta
     4381 tattgtcaat cgcatgcatt agtgaaaata ggcctactac acatataaaa ggcgcgccat
     4441 caacgtcaaa acatgcacac ccaaaaaaaa aaaaaaaaaa aaaaaaaaaa actatcagta
     4501 aaacaaaata cttaatgtac ttacctcaat atgcttcctt aggtttgatg gcgaactttt
     4561 gaaggccgat atttctttat taagcgggag acagaggaca catttcatct tgaatgaatc
     4621 tttatttaca ccacttaaag aaaagaattc gcgaagatac ggccacgggt gctcttgatc
     4681 ctgtggctga ttttggactg tgctgctcgc agctgctgat gaatcacata cttcctccat
     4741 tttcttccac tgattgactg ttataatttc cctaatttcc aggtcaaggt gctgtgcatt
     4801 gtggtaatag atgtgacatg acgtcacttc caaaggacca atgaacatgt ctgaccaatt
     4861 tcatataatg tgaaaacgat tttcataggc agaataaata acatttaaat taaactgggc
     4921 atcagcgcaa ttcaattggt ttggtaatag caagggaaaa tagaatgaag tgatctccaa
     4981 aaaataagta ctttttgact gtaaataaaa ttgtaaggag taaaaagtac ttttttttct
     5041 aaaaaaatgt aattaagtaa aagtaaaagt attgattttt aattgtactc aagtaaagta
     5101 aaaatcccca aaaataatac ttaagtacag taatcaagta aaattactca agtactttac
     5161 acctctggtt cttgaccccc taccttcagc aagcccagca gatccactag ttctagagcg
     5221 gccgccaccg cggtggagct ccagcttttg ttccctttag tgagggttaa ttgcgcgctt
     5281 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     5341 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     5401 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     5461 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     5521 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     5581 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     5641 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     5701 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     5761 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     5821 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     5881 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     5941 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     6001 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     6061 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     6121 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     6181 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     6241 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     6301 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     6361 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     6421 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     6481 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     6541 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     6601 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     6661 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     6721 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     6781 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     6841 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     6901 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     6961 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     7021 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     7081 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     7141 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     7201 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     7261 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     7321 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     7381 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtg
//
