LOCUS       sequence-289215-        6728 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsTDRD3-EBMmut.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6728)
  AUTHORS   Kashima I, Jonas S, Jayachandran U, Buchwald G, Conti E, Lupas AN, 
            Izaurralde E
  TITLE     SMG6 interacts with the exon junction complex via two conserved 
            EJC-binding motifs (EBMs) required for nonsense-mediated mRNA decay.
  JOURNAL   Genes Dev. 2010 Nov 1. 24(21):2440-50.
  PUBMED    20930030
REFERENCE   2  (bases 1 to 6728)
  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: "Genes Dev. 
            2010 Nov 1. 24(21):2440-50."
COMMENT     Sequence Label: pT7-EGFP-C1-HsTDRD3-EBMmut_K
FEATURES             Location/Qualifiers
     source          1..6728
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     613..632
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        613..631
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      659..668
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(665..667,668..670,671..1381)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(665..667,668..670,671..1381)
                     /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(710..731)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(971..990)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1318..1339
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1604..1615
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     polyA_signal    3516..3637
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3553..3572)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3607..3626
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3644..4099)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3781..3802)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3993..4012
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4126..4230
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4232..4589
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4232..4423
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4255..4275)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4440..4575
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4624..5418
                     /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             4624..5418
                     /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(4678..4697)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5288..5307
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5606..5625)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5650..5697
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6026..6614
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6515..6534
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca ctgcttactg
      601 gcttatcgaa attaatacga ctcactatag ggagacccaa gctctagcgc taccggtcgc
      661 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
      721 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
      781 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
      841 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
      901 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
      961 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     1021 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1081 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     1141 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     1201 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     1261 ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat
     1321 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     1381 gtccggactc agatctcgag ctatgctgcg attacagatg actgatggtc atataagttg
     1441 cacagcagta gaatttagtt atatgtcaaa aataagcctg aacacaccac ctggaactaa
     1501 agttaagctc tcaggcattg ttgacataaa aaatggattc ctgctcttga atgactctaa
     1561 caccacagtt cttggtggtg aagtggaaca ccttattgag aaatgggagt tacagagaag
     1621 cttatcaaaa cacaatagaa gcaatattgg aactgaaggt ggaccaccgc cttttgtgcc
     1681 ttttggacag tgtgtatctc gtgtccaagt ggatagcaga gaacttgatc gaagaaaaac
     1741 attgcaagtt acaatgcctg tcaaacctac aaatgataat gatgaatttg aaaagcaaag
     1801 gacggctgct attgctgaag ttgcaaagag caaggaaacc aagacatttg gaggaggtgg
     1861 tggtggtgct agaagtaatc tcaatatgaa tgctgctggt aaccgaaata gggaagtttt
     1921 acagaaagaa aagtcaacca aatcagaggg aaaacatgaa ggtgtctata gagaactggt
     1981 tgatgagaaa gctctgaagc acataacgga aatgggcttc agtaaggaag catcgaggca
     2041 agctcttatg gataatggca acaacttaga agcagcactg aacgtacttc ttacaagcaa
     2101 taaacagaaa cctgttatgg gtcctcctct gagaggtaga ggaaaaggca gggggcgaat
     2161 aagatctgaa gatgaagagg acctgggaaa tgcaaggcca tcagcaccaa gcacattatt
     2221 tgatttcttg gaatctaaaa tgggaacttt gaatgtggaa gaacctaaat cacagccaca
     2281 gcagcttcat cagggacaat acagatcatc aaatactgag caaaatggag taaaagataa
     2341 taatcatctg agacatcctc ctcgaaatga taccaggcag ccaagaaatg aaaaaccgcc
     2401 tcgttttcaa agagactccc aaaattcaaa gtcagtttta gaaggcagtg gattacctag
     2461 aaatagaggt tctgaaagac caagtacttc ttcagtatct gaagtatggg ctgaagacag
     2521 aatcaaatgt gatagaccgt attctagata tgacagaact aaagatactt catatccttt
     2581 aggttctcag catagtgatg gtgcttttaa aaaaagagat aactctatgc aaagcagatc
     2641 aggaaaaggt ccctcctttg cagaggcaaa agaaaatcca cttcctcaag gatctgtaga
     2701 ttataataat caaaaacgtg gaaaaagaga aagccaaaca tctattcctg attattttta
     2761 tgacaggaaa tcacaaacaa taaataatga agctttcagt ggtataaaaa ttgaaaaaca
     2821 ttttaatgta aatactgatt atcagaatcc agttcgaagt aatagtttca ttggtgttcc
     2881 aaatggagaa gtagaaatgc cactgaaagg aagacgaata ggacatatta agccagcagg
     2941 acctgtcaca gctgtaccct gtgatgataa aatattttac aatagtgggc ccaaacgaag
     3001 atctgggcca attaagccag aaaaaatact agaatcatct attcctatgg agtatgcaaa
     3061 aatgtggaaa cctggagatg aatgttttgc actttattgg gaagacaaca agttttaccg
     3121 ggcagaagtt gaagccctcc attcttcggg tatgacagca gttgttaaat tcattgacta
     3181 cggaaactat gaagaggtgc tactgagcaa tatcaagccc attcaaacag aggcatggga
     3241 ggaagaaggc acctacgatc aaactctgga gttccgtagg ggaggtgatg gccagccaag
     3301 acgatccact gaaccaaccc aacagtttga acaaccaccc cgggctcgga actaagaatt
     3361 ctgcagtcga cggtaccgcg ggcccgggat ccaccggatc tagataactg atcataatca
     3421 gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga
     3481 acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg
     3541 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     3601 ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg cgtaaattgt aagcgttaat
     3661 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc
     3721 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt
     3781 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa
     3841 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg
     3901 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
     3961 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
     4021 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
     4081 gcgccgctac agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     4141 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     4201 atgcttcaat aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg
     4261 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     4321 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     4381 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     4441 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     4501 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     4561 gcttttttgg aggcctaggc ttttgcaaag atcgatcaag agacaggatg aggatcgttt
     4621 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     4681 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     4741 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     4801 ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     4861 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     4921 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     4981 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     5041 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     5101 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc
     5161 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     5221 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     5281 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     5341 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     5401 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     5461 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     5521 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     5581 tcgcccaccc tagggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc
     5641 gcgctatgac ggcaataaaa agacagaata aaacgcacgg tgttgggtcg tttgttcata
     5701 aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg
     5761 gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc
     5821 cagggctcgc agccaacgtc ggggcggcag gccctgccat agcctcaggt tactcatata
     5881 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     5941 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     6001 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     6061 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     6121 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     6181 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     6241 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     6301 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     6361 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     6421 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     6481 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     6541 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     6601 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     6661 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     6721 ccatgcat
//
