LOCUS       sequence-330619-        3911 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Standard Gateway entry clone encoding eGFP fluorescent protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3911)
  AUTHORS   Wall VE, Garvey LA, Mehalko JL, Procter LV, Esposito D
  TITLE     Combinatorial assembly of clone libraries using site-specific 
            recombination.
  JOURNAL   Methods Mol Biol. 2014;1116:193-208. doi: 
            10.1007/978-1-62703-764-8_14.
  PUBMED    24395366
REFERENCE   2  (bases 1 to 3911)
  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: "Methods Mol
            Biol."; date: "2014"; volume: "1116:193-208. doi"; pages: " 
            10.1007/978-1-62703-764-8_14"
COMMENT     Sequence Label: C122-E11: eGFP
FEATURES             Location/Qualifiers
     source          1..3911
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      668..677
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(674..676,677..679,680..1393)
                     /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(674..676,677..679,680..1390)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(719..740)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(980..999)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1327..1348
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(1392..1491)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1508..1527)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1509..1527)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1532..1548)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1532..1548)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1979..2770
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     oriT            2892..3001
                     /note="incP origin of transfer"
     rep_origin      3261..3849
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3750..3769
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agttggcgcc accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
      721 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
      781 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
      841 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
      901 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
      961 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     1021 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     1081 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     1141 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     1201 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     1261 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     1321 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     1381 gctgtacaag taatcaactt tcttgtacaa agttggcatt ataagaaagc attgcttatc
     1441 aatttgttgc aacgaacagg tcactatcag tcaaaataaa atcattattt gccatccagc
     1501 tgatatcccc tatagtgagt cgtattacat ggtcatagct gtttcctggc agctctggcc
     1561 cgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat catgcctcct
     1621 ctagaccagc caggacagaa atgcctcgac ttcgctgcta cccaaggttg ccgggtgacg
     1681 cacaccgtgg aaacggatga aggcacgaac ccagtggaca taagcctgtt cggttcgtaa
     1741 gctgtaatgc aagtagcgta tgcgctcacg caactggtcc agaaccttga ccgaacgcag
     1801 cggtggtaac ggcgcagtgg cggttttcat ggcttgttat gactgttttt ttggggtaca
     1861 gtctatgcct cgggcatcca agcagcaagc gcgttacgcc gtgggtcgat gtttgatgtt
     1921 atggagcagc aacgatgtta cgcagcaggg cagtcgccct aaaacaaagt taaacattat
     1981 gagggaagcg gtgatcgccg aagtatcgac tcaactatca gaggtagttg gcgtcatcga
     2041 gcgccatctc gaaccgacgt tgctggccgt acatttgtac ggctccgcag tggatggcgg
     2101 cctgaagcca cacagtgata ttgatttgct ggttacggtg accgtaaggc ttgatgaaac
     2161 aacgcggcga gctttgatca acgacctttt ggaaacttcg gcttcccctg gagagagcga
     2221 gattctccgc gctgtagaag tcaccattgt tgtgcacgac gacatcattc cgtggcgtta
     2281 tccagctaag cgcgaactgc aatttggaga atggcagcgc aatgacattc ttgcaggtat
     2341 cttcgagcca gccacgatcg acattgatct ggctatcttg ctgacaaaag caagagaaca
     2401 tagcgttgcc ttggtaggtc cagcggcgga ggaactcttt gatccggttc ctgaacagga
     2461 tctatttgag gcgctaaatg aaaccttaac gctatggaac tcgccgcccg actgggctgg
     2521 cgatgagcga aatgtagtgc ttacgttgtc ccgcatttgg tacagcgcag taaccggcaa
     2581 aatcgcgccg aaggatgtcg ctgccgactg ggcaatggag cgcctgccgg cccagtatca
     2641 gcccgtcata cttgaagcta gacaggctta tcttggacaa gaagaagatc gcttggcctc
     2701 gcgcgcagat cagttggaag aatttgtcca ctacgtgaaa ggcgagatca ccaaggtagt
     2761 cggcaaataa ccctcgactc tagaggatcc ccgggtaccg agctcgaatt ccgtgatagg
     2821 tgggctgccc ttcctggttg gcttggtttc atcagccatc cgcttgccct catctgttac
     2881 gccggcggta gccggccagc ctcgcagagc aggattcccg ttgagcaccg ccaggtgcga
     2941 ataagggaca gtgaagaagg aacacccgct cgcgggtggg cctacttcac ctatcctgcc
     3001 cggctgacgc cgttggatac accaaggaaa gtctacacga accctttggc aaaatcctgt
     3061 atatcgtgcg aaaaaggatg gatataccga aaaaatcgct ataatgaccc cgaagcaggg
     3121 ttatgcagcg gaaaacggaa ttcgagctcg gtacccgggg atcctctaga gtcgagccac
     3181 ccatgaccaa aatcccttaa cgtgagttac gcgtcgttcc actgagcgtc agaccccgta
     3241 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     3301 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     3361 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     3421 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     3481 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     3541 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     3601 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     3661 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     3721 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     3781 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     3841 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     3901 gctcacatgt t
//
