LOCUS       sequence-203176-        3269 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway Entry vector for EGFP (codon optimized for efficient gene 
            synthesis).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3269)
  AUTHORS   Yamaji M, Jishage M, Meyer C, Suryawanshi H, Der E, Yamaji M, Garzia
            A, Morozov P, Manickavel S, McFarland HL, Roeder RG, Hafner M, 
            Tuschl T
  TITLE     DND1 maintains germline stem cells via recruitment of the CCR4-NOT 
            complex to target mRNAs.
  JOURNAL   Nature. 2017 Mar 23;543(7646):568-572. doi: 10.1038/nature21690. 
            Epub 2017 Mar 15.
  PUBMED    28297718
REFERENCE   2  (bases 1 to 3269)
  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: 
            "10.1038/nature21690"; journalName: "Nature"; date: "2017-03-23- 
            23"; volume: "543"; issue: "7646"; pages: "568-572"
COMMENT     Sequence Label: pENTR4-EGFP_opt
FEATURES             Location/Qualifiers
     source          1..3269
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..100
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             join(106..108,109..111,112..825)
                     /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(106..108,109..111,112..822)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(151..172)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(412..431)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     759..780
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(826..925)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(942..961)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(943..961)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(966..982)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(966..982)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1084..1103)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1095..1904
                     /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(1162..1181)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2051..2639
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2540..2559
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2793..2810
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(2969..2996)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3088..3174)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3223..3245
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3237..3254
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3238..3254
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 caaataatga ttttattttg actgatagtg acctgttcgt tgcaacaaat tgatgagcaa
       61 tgctttttta taatgccaac tttgtacaaa aaagcaggct ccaccatggt gagcaagggc
      121 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
      181 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
      241 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
      301 acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc
      361 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
      421 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
      481 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
      541 tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
      601 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
      661 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag
      721 tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
      781 accgccgccg ggatcactct cggcatggac gagctgtaca agtagaccca gctttcttgt
      841 acaaagttgg cattataaga aagcattgct tatcaatttg ttgcaacgaa caggtcacta
      901 tcagtcaaaa taaaatcatt atttgccatc cagctgatat cccctatagt gagtcgtatt
      961 acatggtcat agctgtttcc tggcagctct ggcccgtgtc tcaaaatctc tgatgttaca
     1021 ttgcacaaga taaaaatata tcatcatgaa caataaaact gtctgcttac ataaacagta
     1081 atacaagggg tgttatgagc catattcaac gggaaacgtc gaggccgcga ttaaattcca
     1141 acatggatgc tgatttatat gggtataaat gggctcgcga taatgtcggg caatcaggtg
     1201 cgacaatcta tcgcttgtat gggaagcccg atgcgccaga gttgtttctg aaacatggca
     1261 aaggtagcgt tgccaatgat gttacagatg agatggtcag actaaactgg ctgacggaat
     1321 ttatgcctct tccgaccatc aagcatttta tccgtactcc tgatgatgca tggttactca
     1381 ccactgcgat ccccggaaaa acagcattcc aggtattaga agaatatcct gattcaggtg
     1441 aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta
     1501 attgtccttt taacagcgat cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata
     1561 acggtttggt tgatgcgagt gattttgatg acgagcgtaa tggctggcct gttgaacaag
     1621 tctggaaaga aatgcataaa cttttgccat tctcaccgga ttcagtcgtc actcatggtg
     1681 atttctcact tgataacctt atttttgacg aggggaaatt aataggttgt attgatgttg
     1741 gacgagtcgg aatcgcagac cgataccagg atcttgccat cctatggaac tgcctcggtg
     1801 agttttctcc ttcattacag aaacggcttt ttcaaaaata tggtattgat aatcctgata
     1861 tgaataaatt gcagtttcat ttgatgctcg atgagttttt ctaatcagaa ttggttaatt
     1921 ggttgtaaca ctggcagagc attacgctga cttgacggga cggcgcaagc tcatgaccaa
     1981 aatcccttaa cgtgagttac gcgtcgttcc actgagcgtc agaccccgta gaaaagatca
     2041 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     2101 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     2161 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     2221 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     2281 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     2341 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     2401 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     2461 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     2521 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     2581 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     2641 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     2701 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     2761 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     2821 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     2881 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat acgcgtaccg
     2941 ctagccagga agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg ccttctgctt
     3001 agtttgatgc ctggcagttt atggcgggcg tcctgcccgc caccctccgg gccgttgctt
     3061 cacaacgttc aaatccgctc ccggcggatt tgtcctactc aggagagcgt tcaccgacaa
     3121 acaacagata aaacgaaagg cccagtcttc cgactgagcc tttcgtttta tttgatgcct
     3181 ggcagttccc tactctcgcg ttaacgctag catggatgtt ttcccagtca cgacgttgta
     3241 aaacgacggc cagtcttaag ctcgggccc
//
