LOCUS       sequence-383068-        7987 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  P2 promoter for transgenic enhancer assays in zebrafish.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7987)
  AUTHORS   Begeman IJ, Emery B, Kurth A, Kang J
  TITLE     Regeneration and developmental enhancers are differentially 
            compatible with minimal promoters.
  JOURNAL   Dev Biol. 2022 Sep 24;492:47-58. doi: 10.1016/j.ydbio.2022.09.007.
  PUBMED    36167150
REFERENCE   2  (bases 1 to 7987)
  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: "Dev Biol.";
            date: "2022-09-24"; volume: "492:47-58. doi"; pages: " 
            10.1016/j.ydbio.2022.09.007"
COMMENT     Sequence Label: Tol2-ISceI-P2:EGFP;cryaa:mCherry-ISceI-Tol2
FEATURES             Location/Qualifiers
     source          1..7987
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(533..1324,1325..1393))
                     /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     1156..1175
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1394..1498)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1524..1979)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1661..1682)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1873..1892
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2106..2128
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2120..2137
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2121..2137
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2147..2165
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(2181..2192,2193..2355)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     CDS             join(4487..4489,4490..4492,4493..5206)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSAQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4532..4553)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4793..4812)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5140..5161
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    5236..5370
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    5377..5498
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(5621..6331)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEDDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGHHSTGGMDELYK"
     primer_bind     complement(5701..5724)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(5899..5918)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     6168..6186
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      6328..6337
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    join(7034..7221,7222..7233)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(7267..7283)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7267..7283)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(7320..7340)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(7320..7338)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(7359..7375)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7359..7375)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7372..7394)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7383..7399
                     /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(7407..7413,7414..7431,7432..7437))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7452..7473
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7590..7607)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(7761..7987,1..362))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7841..7860)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
       61 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
      121 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
      181 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
      241 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
      301 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
      361 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
      421 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
      481 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
      541 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
      601 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
      661 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
      721 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
      781 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
      841 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
      901 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
      961 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     1021 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     1081 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     1141 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     1201 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     1261 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     1321 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     1381 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     1441 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     1501 catttccccg aaaaatgcca cctaaattgt aagcgttaat attttgttaa aattcgcgtt
     1561 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     1621 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     1681 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     1741 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     1801 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     1861 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     1921 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     1981 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     2041 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     2101 gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat acgactcact
     2161 atagggcgaa ttgggtacca cagaggtgta aaaagtactc aaaaatttta ctcaagtgaa
     2221 agtacaagta cttagggaaa attttactca attaaaagta aaagtatctg gctagaatct
     2281 tacttgagta aaagtaaaaa agtactccat taaaattgta cttgagtatt aaggaagtaa
     2341 aagtaaaagc aagaaagaaa actagagatt cttgaagctt tagggataac agggtaatct
     2401 cgagctagcc taggcgccat cgatgtttaa acggatcctg ctcttgaaca agtgactttc
     2461 gttgcaactc tggagatgaa ggggaacaat tattcattta actgacaatt tcaacatgca
     2521 gacatcttat ttattcactt ctttatattg tttgtttgtt tgtacttaat tccatggcat
     2581 ttcattccat atacatctac agttgaagtc agatatatta gcccccctga attattagcc
     2641 cccctattta tttttttccc cattttctgt ttaactgtga gaagattttt ctcaacacat
     2701 ttcttaacat aagttttaat aactcatttc taataactga tttatttgat ctttgccatg
     2761 ttgacagtaa aaaatagaca agatatttga caagacactt ctatgcagct taaggtgaca
     2821 tttaaaggat taatttggtt aattaggtta actaggctac ttagagtaat tagccaagat
     2881 attgattaac gatggtttgt tttggagaga aaaaaaaata gcttaaaggg gctaatcatt
     2941 ttctccttaa aatagttgtt aaaaaattca aaactgcttt tattttttag tcactttgag
     3001 tttactcaaa tgttcttgaa tatttcacat tgttacaatt aacttaaaat gtgcagcaca
     3061 attgtttatt ttaccattta tataactgca taaaacattc cgtagccaat aaagatgtac
     3121 aaataaattg taaactatga accttattac agttaactta gagttgagtc aacacataaa
     3181 acaaatataa catttgctga ctaattattc acatcatttt ctatcaattc tatactggtc
     3241 tatactattg tttctataat atgagatgca ctataagtac tacactctat ctattgtttt
     3301 accacatcat aaatagccac aatcattaat ttagtaaatt ctaacaataa atattacttg
     3361 ggtacaactg gcagataatc tcactaactg tgtatacata tacaaaacaa tacttaagcg
     3421 aacaagccaa tttttgcggt aaaaacttgt gctgttaaac tctgaacaaa ttttaacaaa
     3481 tcaagcccca taaatatccc aactttaatg tattcatatc agcatgacaa ataaagatac
     3541 ggtcctataa aaggtcaata tattgacata tacagcctta aaagtgaaaa caaactccct
     3601 ctaatacatg tggggcccag ctgtcattta tcaagattta ttaaccacaa ctttactgga
     3661 agtgtcctta cctggagccc ccgggtttga tgtttcgggc ttgttgaaag ggtcttctct
     3721 agtcttcaag ttcatgatgg acacaggggt gaaccagacc agaccggcat cagggtgatt
     3781 cactggagaa atgagtcaat catccacagc tataaaaagg gagattcctg cttgagtttc
     3841 tcaatatgca catcaacaca ccagaggtaa gcgctcactc attcttgtca acatgggcac
     3901 caaacatctc tatatgacct ttaggcttct tgactgtgta gtttcattta ttattggact
     3961 gtactcagca acttaagtgc attgattttt ggttttttga attacacaca actaagggac
     4021 tgatttagca tgaatcttca atgtagtgaa gatcataaat cattctaatc gtgctgagtt
     4081 agtggttaaa taaacatatc taattgttga aagcagacat tttagttgaa attatgatta
     4141 tttttaatta ttcttaagtt ttaatgttgg ttttgtattg tctaatgatt aaaatgtata
     4201 aataaaaact aaaaactaaa cataaaaaat agtattactt tcattttgtc taaggaatgt
     4261 caatttgtat ttatttattt attttaaagt gcacaaaata aaaatacatg tgtttgagta
     4321 aaaccttatt taaagtgtat ttaactgttg tgtaaatgag taatgtttag gctagcttta
     4381 ttcattattt tactgtattt aacatattga actgggatat tctgatgtac acaacaaact
     4441 atttgaaaaa atataatttc atttggtctt ttgcagaaat gaattcatgg tgagcaaggg
     4501 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     4561 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     4621 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct
     4681 gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt
     4741 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     4801 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     4861 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     4921 ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa
     4981 cttcaagatc cgccacaaca tcgaggacgg cagcgcgcag ctcgccgacc actaccagca
     5041 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca
     5101 gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     5161 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtagtcta gaactatagt
     5221 gagtcgtatt acgtagatcc agacatgata agatacattg atgagtttgg acaaaccaca
     5281 actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt
     5341 gtaaccatta taagctgcaa taaacaagtt catatgtaag atacattgat gagtttggac
     5401 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     5461 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     5521 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
     5581 aatgtggtat ggctgattat gatctagagt cgcggccgct ttacttgtac agctcgtcca
     5641 tgccgccggt ggagtggtgg ccctcggcgc gctcgtactg ttccacgatg gtgtagtcct
     5701 cgttgtggga ggtgatgtcc agcttgatgt tgacgttgta ggcgccgggc agctgcacgg
     5761 gcttcttggc cttgtaggtg gtcttgacct cggcgtcgta gtggccgccg tccttcagct
     5821 tcagcctctg cttgatctcg cccttcaggg cgccgtcctc ggggtacatc cgctcggagg
     5881 aggcctccca gcccatggtc ttcttctgca ttacggggcc gtcggagggg aagttggtgc
     5941 cgcgcagctt caccttgtag atgaactcgc cgtcctgcag ggaggagtcc tgggtcacgg
     6001 tcaccacgcc gccgtcctcg aagttcatca cgcgctccca cttgaagccc tcggggaagg
     6061 acagcttcaa gtagtcgggg atgtcggcgg ggtgcttcac gtaggccttg gagccgtaca
     6121 tgaactgagg ggacaggatg tcccaggcga agggcagggg gccgcccttg gtcaccttca
     6181 gcttggcggt ctgggtgccc tcgtaggggc ggccctcgcc ctcgccctcg atctcgaact
     6241 cgtggccgtt cacggagccc tccatgtgca ccttgaagcg catgaactcc ttgatgatgg
     6301 ccatgttgtc atcctcgccc ttgctcacca tggtggcgac cggtaatgtc agacctggta
     6361 actccttact gtaacttatt gcacctgatg aacaaaagat gtgaactgaa cgcaagtagg
     6421 aggctaatag aggctagtac agtacaaaga tccccaaatg atgtgtgccg gccttagtgg
     6481 aggcagaccc ctggttatat acctgtgctg cagaaagagg ggcttacaaa tggatctctc
     6541 tggaatggca ttatctcatg gtgggaacag tgttgtcagc agaatcagcg gggtggactc
     6601 tgacctgaga atgacagtct agcactgaat ggctcagaca atactcttgt ggccaactat
     6661 caccggaaaa taagtgcctg ccaaaacaaa ccagaccaga tgaagctaca tggctgtaga
     6721 acgtcagtat ttaatgtgtt gtggccactt ggttcaccag atttaagagc taattttttt
     6781 aaatatcaca gataatcaat cgcttttttt ttttcttgac tgaaaagctt agactggaaa
     6841 attctttgaa aaagtttgca ctacagaaaa taaatacata acatcctgca ctgaatgcac
     6901 actattaata gaaaagagag cattattgta ggaaatgtat tgtatatgta cattctgtgc
     6961 atttcacaat tttgggcccg tagggataac agggtaatca attggtttgg taatagcaag
     7021 ggaaaataga atgaagtgat ctccaaaaaa taagtacttt ttgactgtaa ataaaattgt
     7081 aaggagtaaa aagtactttt ttttctaaaa aaatgtaatt aagtaaaagt aaaagtattg
     7141 atttttaatt gtactcaagt aaagtaaaaa tccccaaaaa taatacttaa gtacagtaat
     7201 caagtaaaat tactcaagta ctttacacct ctggttcttg accccctacc ttcagcaagc
     7261 ccagcagatc cactagttct agagcggccg ccaccgcggt ggagctccag cttttgttcc
     7321 ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt tcctgtgtga
     7381 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     7441 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     7501 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     7561 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     7621 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     7681 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     7741 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     7801 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     7861 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     7921 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     7981 tcggtgt
//
