LOCUS       Exported                3491 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  EGFP (green) fluorophore with multiple cloning site for use with 
            inducible N-terminal fusions..
ACCESSION   .
VERSION     .
KEYWORDS    p3E-EGFP-NS-MCS
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3491)
  AUTHORS   Don EK, Formella I, Badrock AP, Hall TE, Morsch M, Hortle E, Hogan 
            A, Chow S, Gwee SS, Stoddart JJ, Nicholson G, Chung R, Cole NJ
  TITLE     A Tol2 Gateway-Compatible Toolbox for the Study of the Nervous 
            System and Neurodegenerative Disease.
  JOURNAL   Zebrafish. 2016 Sep 15.
  PUBMED    27631880
REFERENCE   2  (bases 1 to 3491)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3491
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      268..295
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      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    complement(624..748)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             762..1478
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             762..1478
                     /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(807..828)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1068..1087)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1415..1436
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1543..1559
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1544..1560
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(1600..1695)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1732..1751)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1733..1751)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1756..1772)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1756..1772)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1874..1893)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1885..2694
                     /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(1952..1971)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2841..3429
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3330..3349
                     /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 tcgggccctg cagctctaga gctcgaattc tacaggtcac
      601 taataccatc taagtagttg gttcatagtg actgcatatg ttgtgtttta cagtattatg
      661 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt
      721 ctcgttcaac tttcttgtac aaagtggctg ccaccaccgg tatggtgagc aagggcgagg
      781 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
      841 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
      901 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
      961 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     1021 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     1081 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     1141 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     1201 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
     1261 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     1321 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg
     1381 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     1441 ccgccgggat cactctcggc atggacgagc tgtacaaggg cggtggaaga tctgggaatt
     1501 caaggcctct cgagcctcta gattctgcag ccctaggatc cctcgaggtc gacggtatcg
     1561 ataagcttga tatcgaattc cccgggtcta gaactagtgc aactttatta tacaaagttg
     1621 gcattataaa aaagcattgc ttatcaattt gttgcaacga acaggtcact atcagtcaaa
     1681 ataaaatcat tatttggagc tccatggtag cgttaacgcg gccgcgatat cccctatagt
     1741 gagtcgtatt acatggtcat agctgtttcc tggcagctct ggcccgtgtc tcaaaatctc
     1801 tgatgttaca ttgcacaaga taaaaatata tcatcatgaa caataaaact gtctgcttac
     1861 ataaacagta atacaagggg tgttatgagc catattcaac gggaaacgtc gaggccgcga
     1921 ttaaattcca acatggatgc tgatttatat gggtataaat gggctcgcga taatgtcggg
     1981 caatcaggtg cgacaatcta tcgcttgtat gggaagcccg atgcgccaga gttgtttctg
     2041 aaacatggca aaggtagcgt tgccaatgat gttacagatg agatggtcag actaaactgg
     2101 ctgacggaat ttatgcctct tccgaccatc aagcatttta tccgtactcc tgatgatgca
     2161 tggttactca ccactgcgat ccccggaaaa acagcattcc aggtattaga agaatatcct
     2221 gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt
     2281 cctgtttgta attgtccttt taacagcgat cgcgtatttc gtctcgctca ggcgcaatca
     2341 cgaatgaata acggtttggt tgatgcgagt gattttgatg acgagcgtaa tggctggcct
     2401 gttgaacaag tctggaaaga aatgcataaa cttttgccat tctcaccgga ttcagtcgtc
     2461 actcatggtg atttctcact tgataacctt atttttgacg aggggaaatt aataggttgt
     2521 attgatgttg gacgagtcgg aatcgcagac cgataccagg atcttgccat cctatggaac
     2581 tgcctcggtg agttttctcc ttcattacag aaacggcttt ttcaaaaata tggtattgat
     2641 aatcctgata tgaataaatt gcagtttcat ttgatgctcg atgagttttt ctaatcagaa
     2701 ttggttaatt ggttgtaaca ctggcagagc attacgctga cttgacggga cggcgcaagc
     2761 tcatgaccaa aatcccttaa cgtgagttac gcgtcgttcc actgagcgtc agaccccgta
     2821 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     2881 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     2941 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     3001 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     3061 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     3121 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     3181 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gcattgagaa
     3241 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     3301 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     3361 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     3421 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     3481 gctcacatgt t
//
