LOCUS       sequence-289901-        6796 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNot10_25-707.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6796)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 6796)
  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_25-707_S
FEATURES             Location/Qualifiers
     source          1..6796
                     /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    3584..3705
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3621..3640)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3675..3694
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3712..4167)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3849..3870)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4061..4080
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4194..4298
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4300..4657
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4300..4491
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4323..4343)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4508..4643
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4692..5486
                     /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             4692..5486
                     /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(4746..4765)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5356..5375
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5674..5693)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5718..5765
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6094..6682
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6583..6602
                     /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 ctgctacctg aagaacagca
     2761 agatccaaag caggaaaatg gggctaaaaa tagtaatcaa ttaggtggga acacagagag
     2821 cagcgaaagc agtgaaactt gcagcagtaa aagccatgat ggagataaat tcattccagc
     2881 tccaccttct tctccattga gaaaacagga attagaaaac ttaaagtgct ccatacttgc
     2941 ttgcagtgcc tacgtggctc tggctttggg tgataacctc atggctttga atcatgcaga
     3001 taaacttctt cagcagccca agctgtcagg atctcttaag tttttgggac atttatatgc
     3061 tgcagaagcc ctcatctctc tcgacagaat atctgatgcc attactcact tgaacccgga
     3121 gaatgtcact gatgtctcct tagggatctc ttcaaatgag caggaccaag gatcagacaa
     3181 aggtgaaaat gaagcaatgg aatcctctgg taagcgggcc cctcagtgct accccagttc
     3241 cgtcaactct gccaggactg tgatgctgtt caaccttggc agcgcttact gcctgaggag
     3301 cgaatatgac aaagcccgaa agtgtctcca ccaggcggct tcaatgatcc atcctaaaga
     3361 ggtgccccct gaggccatct tgctggcagt ctaccttgaa ctgcagaatg gtaatactca
     3421 gctggcctta cagatcatca aaaggaatca gtaaggatcc accggatcta gataactgat
     3481 cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct
     3541 ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc
     3601 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     3661 actgcattct agttgtggtt tgtccaaact catcaatgta tcttaacgcg taaattgtaa
     3721 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     3781 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     3841 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     3901 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     3961 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     4021 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     4081 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     4141 cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa
     4201 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4261 cctgataaat gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct
     4321 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     4381 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     4441 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     4501 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     4561 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     4621 gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag
     4681 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     4741 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     4801 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     4861 tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     4921 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     4981 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     5041 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     5101 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     5161 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga
     5221 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     5281 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     5341 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     5401 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     5461 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     5521 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     5581 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     5641 ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga
     5701 aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt
     5761 tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc
     5821 ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg
     5881 tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta
     5941 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     6001 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     6061 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     6121 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     6181 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     6241 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     6301 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     6361 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     6421 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     6481 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     6541 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     6601 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     6661 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     6721 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     6781 tattaccgcc atgcat
//
