LOCUS       sequence-385114-        3736 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway p3E 2A fluorophore.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3736)
  AUTHORS   Kemmler CL, Moran HR, Murray BF, Scoresby A, Klem JR, Eckert RL, 
            Lepovsky E, Bertho S, Nieuwenhuize S, Burger S, D'Agati G, Betz C, 
            Puller AC, Felker A, Ditrychova K, Botschi S, Affolter M, Rohner N, 
            Lovely CB, Kwan KM, Burger A, Mosimann C
  TITLE     Next-generation plasmids for transgenesis in zebrafish and beyond.
  JOURNAL   Development. 2023 Mar 28:dev.201531. doi: 10.1242/dev.201531.
  PUBMED    36975217
REFERENCE   2  (bases 1 to 3736)
  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: 
            "Development. 2023 Mar 28:dev.201531. doi: 10.1242/dev.201531."
COMMENT     Sequence Label: 458 p3E-2A-EGFP-CAAXpA
FEATURES             Location/Qualifiers
     source          1..3736
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(88..211)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             223..279
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             join(280..282,283..285,286..996)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(280..282,283..285,286..996)
                     /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(325..346)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(586..605)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     933..954
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1112..1246
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1136..1155)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1190..1209
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    complement(1309..1404)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(1442..1460)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1465..1481)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1465..1481)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1583..1602)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1594..2403
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1661..1680)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2550..3138
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3039..3058
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3292..3309
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3468..3495)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3587..3673)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(3722..3736,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(3736,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtctt aagctcgggc cctgcagctc tagagctcga attctacagg
       61 tcactaatac catctaagta gttggttcat agtgactgca tatgttgtgt tttacagtat
      121 tatgtagtct gttttttatg caaaatctaa tttaatatat tgatatttat atcattttac
      181 gtttctcgtt caactttctt gtacaaagtg gggggatccg gagccacgaa cttctctctg
      241 ttaaagcaag caggagacgt ggaagaaaac cccggtccta tggtgagcaa gggcgaggag
      301 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
      361 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
      421 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
      481 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
      541 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
      601 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
      661 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
      721 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
      781 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
      841 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc
      901 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
      961 gccgggatca ctctcggcat ggacgagctg tacaagggag gaggaagatc taagctgaac
     1021 cctcctgatg agagtggccc cggctgcatg agctgcaagt gtgtgctctc ctgaggcgcg
     1081 cctctagaac tatagtgagt cgtattacgt agatccagac atgataagat acattgatga
     1141 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     1201 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     1261 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttcca actttattat
     1321 acaaagttgg cattataaaa aagcattgct tatcaatttg ttgcaacgaa caggtcacta
     1381 tcagtcaaaa taaaatcatt atttggagct ccatggtagc gttaacgcgg ccgcgatatc
     1441 ccctatagtg agtcgtatta catggtcata gctgtttcct ggcagctctg gcccgtgtct
     1501 caaaatctct gatgttacat tgcacaagat aaaaatatat catcatgaac aataaaactg
     1561 tctgcttaca taaacagtaa tacaaggggt gttatgagcc atattcaacg ggaaacgtcg
     1621 aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg ggctcgcgat
     1681 aatgtcgggc aatcaggtgc gacaatctat cgcttgtatg ggaagcccga tgcgccagag
     1741 ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga gatggtcaga
     1801 ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat ccgtactcct
     1861 gatgatgcat ggttactcac cactgcgatc cccggaaaaa cagcattcca ggtattagaa
     1921 gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct gcgccggttg
     1981 cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg tctcgctcag
     2041 gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg attttgatga cgagcgtaat
     2101 ggctggcctg ttgaacaagt ctggaaagaa atgcataaac ttttgccatt ctcaccggat
     2161 tcagtcgtca ctcatggtga tttctcactt gataacctta tttttgacga ggggaaatta
     2221 ataggttgta ttgatgttgg acgagtcgga atcgcagacc gataccagga tcttgccatc
     2281 ctatggaact gcctcggtga gttttctcct tcattacaga aacggctttt tcaaaaatat
     2341 ggtattgata atcctgatat gaataaattg cagtttcatt tgatgctcga tgagtttttc
     2401 taatcagaat tggttaattg gttgtaacac tggcagagca ttacgctgac ttgacgggac
     2461 ggcgcaagct catgaccaaa atcccttaac gtgagttacg cgtcgttcca ctgagcgtca
     2521 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     2581 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     2641 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     2701 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     2761 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     2821 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     2881 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     2941 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     3001 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     3061 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     3121 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     3181 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     3241 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     3301 gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg
     3361 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac
     3421 gcaattaata cgcgtaccgc tagccaggaa gagtttgtag aaacgcaaaa aggccatccg
     3481 tcaggatggc cttctgctta gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc
     3541 accctccggg ccgttgcttc acaacgttca aatccgctcc cggcggattt gtcctactca
     3601 ggagagcgtt caccgacaaa caacagataa aacgaaaggc ccagtcttcc gactgagcct
     3661 ttcgttttat ttgatgcctg gcagttccct actctcgcgt taacgctagc atggatgttt
     3721 tcccagtcac gacgtt
//
