LOCUS       sequence-288585-        6127 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of tubcDNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6127)
  AUTHORS   Wittkopp N, Huntzinger E, Weiler C, Sauliere J, Schmidt S, Sonawane 
            M, Izaurralde E
  TITLE     Nonsense-mediated mRNA decay effectors are essential for zebrafish 
            embryonic development and survival.
  JOURNAL   Mol Cell Biol. 2009 Jul . 29(13):3517-28.
  PUBMED    19414594
REFERENCE   2  (bases 1 to 6127)
  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: "Mol Cell 
            Biol. 2009 Jul . 29(13):3517-28."
COMMENT     Sequence Label: pT7-EGFP-C1-tubcDNA_D
FEATURES             Location/Qualifiers
     source          1..6127
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        9..312
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        313..516
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     467..487
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     561..580
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        561..579
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /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(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2863..2984
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2900..2919)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2954..2973
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2991..3446)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3128..3149)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3340..3359
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3473..3577
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3579..3936
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3579..3770
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3602..3622)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3787..3922
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3971..4765
                     /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             3971..4765
                     /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(4025..4044)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4635..4654
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4953..4972)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4997..5044
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5373..5961
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5862..5881
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
       61 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
      121 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
      181 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
      241 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
      301 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
      361 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
      421 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
      481 aggcgtgtac ggtgggaggt ctatataagc agagctctct ggctaactag agaacccact
      541 gcttactggc ttatcgaaat taatacgact cactataggg agacccaagc tctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct caagcttcga attctgcagt cgacggtacc
     1381 atgagggaga tcgtgcattt acaggctgga cagtgcggca accagattgg tgccaagttc
     1441 tgggaagtca ttagtgacga gcatggaatt gacccaacag gcagttacca tggcgacagt
     1501 gaccttcagc tggaccgaat taatgtttat tataatgaag ccacaggtgg aaagtatgtt
     1561 ccacgtgccg tgctggtgga tttggagcct ggtacaatgg actccgtgag gtctggtcca
     1621 tttggtcaga tcttcagacc agacaacttt gtgttcggcc aaagtggtgc tggaaacaac
     1681 tgggccaagg gccactacac tgaaggagct gagctggttg attccgttct ggatgtggtc
     1741 cgaaaagagg ctgagagctg cgactgtctg cagggcttcc agctcactca ctcactgggt
     1801 ggaggtacag ggtctggtat gggcaccctc ctcattagca aaatccgcga ggagtatccc
     1861 gaccgcatca tgaacacctt cagcgtggtg ccctctccta aagtctcgga cactgtggtc
     1921 gagccctaca acgccacact gtccgtccat cagctagtag agaacacaga cgagacctat
     1981 tgtattgata acgaggccct gtacgatatc tgcttccgca cactcaaact cacaaccccc
     2041 acatacggag acctcaacca tctcgtctcc gccacaatga gcggtgtgac cacttgcttg
     2101 aggtttccag gccagttgaa cgctgatctc cgtaaattgg cggtcaacat ggtgcccttc
     2161 ccccgactgc acttcttcat gcctggcttc gcgcctctga ctagcagggg aagccagcag
     2221 tatcgtgcac ttaccgttcc cgaactcacc cagcagatgt tcgatgccaa aaacatgatg
     2281 gctgcctgcg acccacgtca cggccgttat ctgacggtcg ccgccgtctt ccgaggtcgc
     2341 atgtccatga aggaggtgga cgagcagatg ctcaacgtcc agaacaagaa cagcagctac
     2401 ttcgttgaat ggatcccaaa caacgtcaag accgccgtct gcgacattcc accacgaggc
     2461 ctcaagatgg ccgccacctt catcggcaac agcactgcca tccaggagct cttcaagcgc
     2521 atctcagagc agttcacggc tatgttcagg cgcaaggctt tcctgcattg gtacaccgga
     2581 gagggcatgg atgagatgga gttcacagag gccgagagca acatgaacga cctggtgtcc
     2641 gagtaccagc agtaccagga cgccaccgct gaagaggaag gggagtttga ggaggaggga
     2701 gaaaaggagc tcgcttaagg taccgcgggc ccgggatcca ccggatctag ataactgatc
     2761 ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc
     2821 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct
     2881 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     2941 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaacgcgt aaattgtaag
     3001 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     3061 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     3121 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     3181 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     3241 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     3301 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     3361 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     3421 gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac
     3481 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     3541 ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg
     3601 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3661 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     3721 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     3781 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     3841 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     3901 tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg
     3961 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     4021 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     4081 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     4141 gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     4201 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     4261 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     4321 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     4381 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     4441 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag
     4501 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     4561 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     4621 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     4681 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     4741 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     4801 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     4861 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     4921 gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa
     4981 ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt
     5041 gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc
     5101 cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt
     5161 gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac
     5221 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     5281 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     5341 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     5401 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     5461 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     5521 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     5581 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     5641 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     5701 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     5761 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     5821 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     5881 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     5941 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     6001 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     6061 attaccgcca tgcattagtt attaatagta atcaattacg gggtcattag ttcatagccc
     6121 atatatg
//
