LOCUS       sequence-289213-        8669 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsRRP12-EBMmut.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8669)
  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 8669)
  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-HsRRP12-EBMmut_K
FEATURES             Location/Qualifiers
     source          1..8669
                     /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"
     polyA_signal    5457..5578
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5494..5513)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5548..5567
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5585..6040)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5722..5743)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5934..5953
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6067..6171
                     /gene="bla"
                     /label=AmpR promoter
     promoter        6173..6530
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6173..6364
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6196..6216)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6381..6516
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6565..7359
                     /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             6565..7359
                     /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(6619..6638)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7229..7248
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7547..7566)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7591..7638
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7967..8555
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8456..8475
                     /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 ctatgggtcg ctcgggaaag ttgccctctg gtgtctcagc
     1441 taagttgaag cgctggaaga aaggccacag cagcgacagc aaccccgcca tctgccgcca
     1501 ccgtcaggcc gcccgcagcc gcttcttcag ccggccgtca ggaaggagtg acctgacagt
     1561 cgatgctgtg aagttacata atgagctgca gtcagggtcc ttgcgcttgg gcaaaagcga
     1621 agccccggag acgcccatgg aagaagaggc ggagctggtt ctcaccgaga agtcctcggg
     1681 taccttcctg agtggccttt ccgactgcac aaacgtcacc ttcagcaaag tacagcgctt
     1741 ctgggagtcc aactcggctg cccacaagga gatctgtgct gttctggctg ctgtcactga
     1801 ggtgattcgc tcccagggag ggaaggagac ggagactgag tacttcgctg ctctgatgac
     1861 aacaatggaa gcagtggagt ccccggagtc cctggccgcc gttgcttacc tgctgaacct
     1921 tgtcctgaag cgtgttccca gccctgtgct tattaagaag ttctctgata cctccaaagc
     1981 cttcatggat atcatgtcag ctcaggccag cagcggctcc acctctgtcc tccgatgggt
     2041 cctttcctgc ctggccaccc ttctgcggaa gcaagacctg gaggcctggg gctaccccgt
     2101 gacccttcag gtgtaccatg ggctgctgag cttcacggtg catcccaagc ccaagatccg
     2161 gaaggctgcc cagcatggag tatgctcagt cctcaagggc agtgaattca tgtttgaaaa
     2221 ggcccctgcc catcatcctg ctgccatttc cactgccaag ttctgcatcc aggagattga
     2281 gaagtctgga ggctccaagg aggccaccac cacgctgcac atgctgacgc tgctgaagga
     2341 cctgctgccc tgcttcccgg aaggcctggt gaagagctgc agtgagactc tcctcagggt
     2401 catgaccttg agccatgtgc tggtgacagc ctgtgccatg caggcctttc acagcctctt
     2461 ccacgccagg cctggcctga gcaccctgtc agcagagctc aacgcccaga tcatcacggc
     2521 cctgtacgac tatgttccca gtgagaatga tttacaaccc ctgctagcct ggcttaaggt
     2581 catggagaaa gcccacatca acctggtgag gttgcagtgg gacctggggc taggccacct
     2641 ccctcgcttt tttggaactg cggtgacctg cctcctttcc ccacactcgc aagtgctgac
     2701 tgctgctacg cagagcctca aggagatcct gaaggaatgc gtggctcccc acatggctga
     2761 cattggctcc gtgacctcct cggcctcagg ccctgcccaa tctgttgcca agatgttcag
     2821 ggcagtggag gagggcctga cgtacaaatt ccatgcggcc tggagctccg tgttgcagct
     2881 gctgtgtgtc ttcttcgagg cgtgtgggag acaggcccac cctgtgatga ggaagtgcct
     2941 ccagtccctg tgtgacctgc gcctctcccc tcatttcccc cacacggcgg ctcttgacca
     3001 ggcagtgggg gctgcggtga ccagtatggg acctgaggtg gtgctgcagg ctgtgccttt
     3061 ggaaattgat ggctctgagg agactctgga tttcccacgg agctggctgc tgcctgtcat
     3121 ccgagaccat gttcaggaaa cgcgacttgg ttttttcacc acctacttct tgcccctggc
     3181 taacaccctg aagagcaaag ccatggacct ggctcaggca ggcagcacag tggaatctaa
     3241 gatctacgac acactccagt ggcagatgtg gacactcctg cctgggttct gcacaaggcc
     3301 tacagatgtg gccatctcct tcaaagggct ggcacggacg ctgggcatgg ccatcagcga
     3361 gcgtccagac ctgagggtca ccgtgtgcca ggccctgcgc accctcatca ccaagggctg
     3421 ccaggcagag gctgaccgtg ctgaagtgag tcgctttgcc aagaactttc tgccgatcct
     3481 cttcaacctg tatgggcagc ccgtggcagc cggggacact ccagcccctc gccgggctgt
     3541 gctggaaacc atcagaactt acctcaccat cactgacact cagttggtga acagtctcct
     3601 ggaaaaagcc agtgagaagg tgctcgaccc tgccagctct gactttacca gattgtctgt
     3661 cctggacctg gtcgtggcct tggctccgtg tgctgacgaa gctgccatca gtaagctata
     3721 ctccaccatc cggccctacc tagagagcaa ggcccacggg gtgcagaaga aggcctaccg
     3781 agtgctggag gaggtgtgtg ccagtcctca gggccccggg gccctcttcg tgcagagcca
     3841 cctggaggac ctgaagaaga cactgctgga ctcgctgcgg agcacctcct cacccgccaa
     3901 gaggccccgt ttgaagtgcc tcctacacat cgtgaggaag ctctcagctg aacacaagga
     3961 gttcatcact gccctcatcc cagaggtgat cctgtgcacc aaggaggtgt cggtgggcgc
     4021 acggaagaac gcttttgcac tgctcgtgga gatgggccat gctttcctaa ggtttggctc
     4081 gaaccaggaa gaggccctgc agtgctacct cgtcctgatc taccctggcc tggtgggcgc
     4141 ggtgaccatg gtcagctgca gcatcctggc cctgacccac ctccttttcg agtttaaagg
     4201 tctgatgggg accagtacag tggagcagct gctggagaat gtgtgcctgc ttctggcctc
     4261 ccgcacccgt gacgtggtca agtctgcact gggcttcatc aaggtggcag tgactgtcat
     4321 ggacgtggcg cacctggcca aacatgtgca gctggtgatg gaagccattg ggaagctttc
     4381 agatgacatg cggcggcact tccgcatgaa gcttcggaac ctgttcacca agttcatccg
     4441 caagtttgga tttgagctgg tgaaaaggct gttgcccgag gagtaccaca gagtcctggt
     4501 caacatccgg aaagctgagg cccgggccaa gaggcaccga gccctgagcc aggctgccgt
     4561 ggaggaggaa gaagaggagg aggaggagga ggagcccgcc cagggcaaag gtgacagcat
     4621 tgaggagatt ttagctgact cagaggacga ggaggacaat gaggaggagg aaagaagccg
     4681 aggcaaggag cagcggaagc tggcacgaca gaggagccgg gcatggctga aagagggcgg
     4741 tggggacgag cccctcaact tcctggatcc caaggtggcc caacgagtcc tggccacgca
     4801 gccagggcca ggccggggca ggaagaagga ccacagcttc aaggtgagcg ccgatggccg
     4861 gctgatcata agggaggagg cagacggcaa caagatggag gaagaggaag gtgccaaagg
     4921 cgaagatgaa gagatggctg acccaatgga agatgtgatc atcaggaata aaaagcacca
     4981 gaagctcaag caccagaaag aggctgagga ggaggagctg gagatacccc ctcagtacca
     5041 agctggaggc tctggcattc atcgccctgt ggccaagaag gctatgcctg gggctgaata
     5101 caaggccaag aaagcaaaag gtgatgtgaa gaagaaaggc gaaccggatc cctatgccga
     5161 aatccccctc aacagaagca agctcaaccg caggaagaag atgaagctgc agggacagtt
     5221 caaaggcctg gtgaaggctg cccggcgagg ttcccaggtg ggacacaaaa accgcagaaa
     5281 ggatcgtcga ccctgacaat tctgcagtcg acggtaccgc gggcccggga tccaccggat
     5341 ctagataact gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa
     5401 acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact
     5461 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     5521 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaac
     5581 gcgtaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     5641 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     5701 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     5761 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     5821 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     5881 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     5941 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     6001 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac ttttcgggga
     6061 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     6121 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tcctgaggcg
     6181 gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     6241 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     6301 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     6361 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     6421 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     6481 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa
     6541 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     6601 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     6661 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     6721 ctgtccggtg ccctgaatga actgcaagac gaggcagcgc ggctatcgtg gctggccacg
     6781 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     6841 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     6901 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     6961 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     7021 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     7081 aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     7141 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     7201 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     7261 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     7321 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     7381 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     7441 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     7501 gggatctcat gctggagttc ttcgcccacc ctagggggag gctaactgaa acacggaagg
     7561 agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat aaaacgcacg
     7621 gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac tctgtcgata
     7681 ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc ccaccccacc
     7741 ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca ggccctgcca
     7801 tagcctcagg ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     7861 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     7921 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     7981 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     8041 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     8101 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     8161 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     8221 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     8281 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     8341 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     8401 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     8461 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     8521 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     8581 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     8641 ctgtggataa ccgtattacc gccatgcat
//
