LOCUS       sequence-414170-        4867 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4867)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 4867)
  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"
COMMENT     Sequence Label: pT7-lambdaN-EGFP-C1_Q
FEATURES             Location/Qualifiers
     source          1..4867
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     553..572
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        553..571
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      599..608
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             605..670
                     /codon_start=1
                     /product="N-terminal RNA-binding domain from the lambda 
                     bacteriophage antiterminator protein N (Baron-Benhamou et 
                     al., 2004)"
                     /label=lambda N peptide
                     /translation="MDAQTRRRERRAEKQAQWKAAN"
     CDS             join(689..691,692..694,695..1405)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(689..691,692..694,695..1405)
                     /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(734..755)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(995..1014)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1342..1363
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1406..1471
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    1595..1716
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1632..1651)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1686..1705
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1723..2178)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1860..1881)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2072..2091
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2205..2309
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2311..2668
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2311..2502
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2334..2354)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2519..2654
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2703..3497
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2703..3497
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2757..2776)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3367..3386
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3685..3704)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3729..3776
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4105..4693
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4594..4613
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca ctgcttactg
      541 gcttatcgaa attaatacga ctcactatag ggagacccaa gctctagcgc taccggtcgc
      601 caccatggac gcacaaacac gacgacgtga gcgtcgcgct gagaaacaag ctcaatggaa
      661 agctgcaaac ggtggatctc ctccacatat ggtgagcaag ggcgaggagc tgttcaccgg
      721 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
      781 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
      841 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
      901 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
      961 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     1021 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     1081 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
     1141 ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa
     1201 catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
     1261 cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga
     1321 ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac
     1381 tctcggcatg gacgagctgt acaagtccgg actcagatct cgagctcaag cttcgaattc
     1441 tgcagtcgac ggtaccgcgg gcccgggatc caccggatct agataactga tcataatcag
     1501 ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa
     1561 cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg
     1621 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     1681 tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta agcgttaata
     1741 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     1801 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     1861 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     1921 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     1981 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     2041 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     2101 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     2161 cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     2221 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     2281 tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt
     2341 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     2401 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     2461 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     2521 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     2581 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
     2641 cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc
     2701 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     2761 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     2821 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     2881 tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     2941 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     3001 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     3061 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     3121 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     3181 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg
     3241 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     3301 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     3361 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     3421 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     3481 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     3541 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     3601 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     3661 cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg
     3721 cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa
     3781 acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg
     3841 ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc
     3901 agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat
     3961 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     4021 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     4081 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     4141 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     4201 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     4261 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     4321 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     4381 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     4441 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     4501 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     4561 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     4621 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     4681 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     4741 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     4801 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
     4861 agttccg
//
