LOCUS       sequence-193550-        5338 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent vector for screening enhancer elements in zebrafish.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5338)
  AUTHORS   Chong-Morrison V, Mayes S, Simoes FC, Senanayake U, Carroll DS, 
            Riley PR, Wilson SW, Sauka-Spengler T
  TITLE     Ac/Ds transposition for CRISPR/dCas9-SID4x epigenome modulation in 
            zebrafish.
  JOURNAL   Biol Open. 2023 Jun 15;12(6):bio059995. doi: 10.1242/bio.059995. 
            Epub 2023 Jun 27.
  PUBMED    37367831
REFERENCE   2  (bases 1 to 5338)
  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: "Biol 
            Open."; date: "2023-06-15"; volume: "12(6):bio059995. doi"; pages: "
            10.1242/bio.059995. Epub 2023 Jun 27"
COMMENT     Sequence Label: pVC-Ds-E1b:eGFP-Ds
FEATURES             Location/Qualifiers
     source          1..5338
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(33..461)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(143..164)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     355..374
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     588..610
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     625..644
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        625..643
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          1051..1623
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     regulatory      1697..1706
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1703..1705,1706..1708,1709..2422)
                     /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(1703..1705,1706..1708,1709..2419)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1748..1769)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2009..2028)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2356..2377
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2637..2718
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2674..2693)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2728..2747
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        complement(3153..3171)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3192..3208)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3192..3208)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3205..3227)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3216..3232
                     /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(3240..3246,3247..3264,3265..3270))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3285..3306
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3423..3440)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3594..4182)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3674..3693)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4353..5144,5145..5213))
                     /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     4976..4995
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5214..5318)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
       61 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
      121 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
      181 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
      241 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
      301 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
      361 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
      421 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
      481 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
      541 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
      601 ttgtaaaacg acggccagcg cgcgtaatac gactcactat agggcgaatt gggtacccga
      661 cgcagggatg aaagtaggat gggaaaatcc cgtaccgacc gttatcgtat aaccgatttt
      721 gttagtttta tcccgatcga tttcgaaccc gaggtaaaaa acgaaaacgg aacggaaacg
      781 ggatatacaa aacggtaaac ggaaacggaa acggtagagc tagtttcccg accgtttcac
      841 cgggatcccg tttttaatcg ggatgatccc gtttcgttac cgtattttct aattcgggat
      901 gactgcaaca gatccctcga ctcgagttta cgtaccgcta gcaagcttgc ggcgatatcg
      961 actctagagg gtatataatg gatcccatcg cgtctcagcc tcactttgag ctcctccaca
     1021 cgaatttcga ccgatcctga gaacttcagg gtgagtttgg ggacccttga ttgttctttc
     1081 tttttcgcta ttgtaaaatt catgttatat ggagggggca aagttttcag ggtgttgttt
     1141 agaatgggaa gatgtccctt gtatcaccat ggaccctcat gataattttg tttctttcac
     1201 tttctactct gttgacaacc attgtctcct cttattttct tttcattttc tgtaactttt
     1261 tcgttaaact ttagcttgca tttgtaacga atttttaaat tcacttttgt ttatttgtca
     1321 gattgtaagt actttctcta atcacttttt tttcaaggca atcagggtat attatattgt
     1381 acttcagcac agttttagag aacaattgtt ataattaaat gataaggtag aatatttctg
     1441 catataaatt ctggctggcg tggaaatatt cttattggta gaaacaacta caccctggtc
     1501 atcatcctgc ctttctcttt atggttacaa tgatatacac tgtttgagat gaggataaaa
     1561 tactctgagt ccaaaccggg cccctctgct aaccatgttc atgccttctt ctctttccta
     1621 cagctcctgg gcaacgtgct ggttgttgtg ctgtctcatc attttggcaa agaattcctc
     1681 gacggatcca ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt
     1741 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
     1801 gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa
     1861 gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag
     1921 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
     1981 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
     2041 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     2101 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
     2161 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
     2221 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     2281 cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa
     2341 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
     2401 catggacgag ctgtacaagt aaagcggccg cgactctaga attccaactg agcgccggtc
     2461 gctaccatta ccaacttgtc tggtgtcaaa aataataggc ctactagtga tccaccggat
     2521 ctagataact gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa
     2581 acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact
     2641 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     2701 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtagagtaga
     2761 cttatatggc ttcttatgtt agccaagagc ccaagactta tcacttatgt gctacattaa
     2821 actatgtgtg ctccagattt atatggattt tatctatgtt taattaagac ttgtgtttac
     2881 aattttttat atttgttttt aagttttgaa tatatgtttt catgtgtgat tttaccgaac
     2941 aaaaataccg gttcccgtcc gatttcgact ttaacccgac cggatcgtat cggttttcga
     3001 ttaccgtatt tatcccgttc gttttcgtta ccggtatatc ccgttttcgt ttccgtcccg
     3061 caagttaaat atgaaaatga aaacggtaga ggtattttac cgaccgttac cgaccgtttt
     3121 catccctagt cggggagctc cagcttttgt tccctttagt gagggttaat ttcgagcttg
     3181 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3241 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     3301 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3361 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     3421 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3481 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     3541 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3601 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     3661 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3721 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     3781 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     3841 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     3901 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     3961 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4021 ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4081 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4141 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4201 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4261 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4321 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4381 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4441 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4501 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4561 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4621 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     4681 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     4741 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     4801 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     4861 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     4921 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     4981 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5041 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5101 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5161 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5221 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     5281 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgc
//
