LOCUS       sequence-289900-        6088 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNot10_25-471.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6088)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 6088)
  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-EGFP-C1-HsNot10_25-471_S
FEATURES             Location/Qualifiers
     source          1..6088
                     /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    2876..2997
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2913..2932)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2967..2986
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3004..3459)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3141..3162)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3353..3372
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3486..3590
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3592..3949
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3592..3783
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3615..3635)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3800..3935
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3984..4778
                     /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             3984..4778
                     /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(4038..4057)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4648..4667
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4966..4985)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5010..5057
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5386..5974
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5875..5894
                     /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 ctgatcaaga gaaggagtta tccaccaatg ctttccaagc
     1441 tttcacatct ggaaattatg atgcctgtct acaacacctt gcctgtctac aagatataaa
     1501 caaagatgat tataaaataa ttttgaatac agcagtagct gagtttttta aaagtaacca
     1561 aacaacaaca gataatttga gacaaacact taaccagctg aagaatcagg tccactcagc
     1621 tgttgaagaa atggatggat tagatgatgt tgaaaacagc atgttgtact ataatcaagc
     1681 agtcattctt tatcatctgc ggcagtatac agaagccata tcagttggtg aaaaacttta
     1741 tcagttcata gagccttttg aagaaaaatt tgcccaagca gtgtgttttt tgcttgtaga
     1801 cctgtatata ttaacctacc aagctgagaa agctttgcat cttcttgctg tcctagaaaa
     1861 aatgatttca cagggtaaca ataacaaaaa tggaaagaat gagactggta ataacaacaa
     1921 caaagatgga tctaatcata aagctgaaag tggagctcta atagaagctg caaaatcaaa
     1981 gatacatcag tacaaagtac gagcttatat ccaaatgaag tctctgaaag catgtaaaag
     2041 ggaaatcaag tcagtcatga atacagctgg aaattccgca ccctctctct ttcttaaaag
     2101 caattttgag tacttaagag gtaattatcg aaaagccgtg aagctattaa atagttcaaa
     2161 cattgctgag catccaggat tcatgaaaac aggtgaatgc ttgagatgca tgttctggaa
     2221 taaccttggt tgcatccatt ttgccatgag caagcacaat ttgggaatat tctactttaa
     2281 aaaggctctg caagagaatg acaatgtctg tgcacagctc agtgcaggta gcactgatcc
     2341 aggtaaaaaa ttttcaggaa gacccatgtg tacgttacta accaataaga gatatgagtt
     2401 gctgtataac tgtggaattc agcttcttca cattggaagg cctcttgctg ccttcgaatg
     2461 tctgattgaa gctgttcagg tttatcatgc aaatcctcgc ctctggctac ggctggctga
     2521 atgctgcatt gctgccaata aggggacttc tgaacaagaa actaaaggcc ttcccagcaa
     2581 aaaaggaatt gtacagtcta ttgttggtca aggctatcat cgtaaaatag ttttggcatc
     2641 acagtctata cagaatactg tttataatga tggtcagtct tcggccattc ctgtagccag
     2701 tatggagttt gcagccatat gtctcagaaa tgccttgttg ctgtaaggat ccaccggatc
     2761 tagataactg atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa
     2821 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt
     2881 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     2941 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg
     3001 cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     3061 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     3121 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     3181 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     3241 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     3301 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     3361 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     3421 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa
     3481 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     3541 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg
     3601 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     3661 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     3721 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     3781 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     3841 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     3901 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag
     3961 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     4021 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     4081 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     4141 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga
     4201 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     4261 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     4321 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     4381 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     4441 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     4501 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     4561 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     4621 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     4681 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     4741 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     4801 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     4861 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     4921 ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga
     4981 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg
     5041 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac
     5101 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc
     5161 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat
     5221 agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     5281 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     5341 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     5401 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     5461 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     5521 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     5581 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     5641 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     5701 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     5761 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     5821 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     5881 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     5941 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     6001 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     6061 tgtggataac cgtattaccg ccatgcat
//
