LOCUS       sequence-289184-        5344 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNot6_1-188.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5344)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 5344)
  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-HsNot6_1-188_K
FEATURES             Location/Qualifiers
     source          1..5344
                     /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    2132..2253
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2169..2188)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2223..2242
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2260..2715)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2397..2418)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2609..2628
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2742..2846
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2848..3205
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2848..3039
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2871..2891)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3056..3191
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3240..4034
                     /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             3240..4034
                     /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(3294..3313)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3904..3923
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4222..4241)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4266..4313
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4642..5230
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5131..5150
                     /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 ctcaagcttc gaattctgca gtcgacggta ccgcgggccc
     1441 caaagaaaaa tacgagcccc ctgaccctcg gaggatgtat acaattatgt cttctgagga
     1501 agcagcaaat ggaaagaaat cccactgggc agagcttgaa ataagtggaa aagtaagaag
     1561 cttaagcgca tctttgtggt cactaactca cctgacagct ttgcatttga gtgacaattc
     1621 cctgtcccga attccttcag acattgccaa gcttcacaat ctggtgtatt tggacctgtc
     1681 atctaataaa attcgtagct tacccgcaga actcggaaac atggtatcac tcagggagct
     1741 ccatttaaat aacaacctgt tacgagttct accttttgag ctgggaaaac tgtttcagtt
     1801 gcagacttta ggcctgaaag gaaatcccct tacccaggat atattgaacc tttatcagga
     1861 accagatgga acaagacggc tgctgaacta tttgcttgat aatttgtcag gtactgcaaa
     1921 aagaattaca acagaacaac cacctccaag gtcttggatt atgttacaag aaccagatag
     1981 gacaaggcca actgccttgt agggatccac cggatctaga taactgatca taatcagcca
     2041 taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct
     2101 gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta
     2161 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     2221 ttgtggtttg tccaaactca tcaatgtatc ttaacgcgta aattgtaagc gttaatattt
     2281 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     2341 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     2401 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     2461 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     2521 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     2581 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     2641 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     2701 cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     2761 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     2821 ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt ggaatgtgtg
     2881 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     2941 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3001 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     3061 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     3121 ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt
     3181 ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga tcgtttcgca
     3241 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     3301 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     3361 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     3421 aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     3481 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     3541 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     3601 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     3661 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     3721 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc atgcccgacg
     3781 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     3841 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     3901 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     3961 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     4021 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct
     4081 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     4141 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     4201 ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag gaacccgcgc
     4261 tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg ttcataaacg
     4321 cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc attggggcca
     4381 atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg aaggcccagg
     4441 gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact catatatact
     4501 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     4561 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     4621 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     4681 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     4741 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta
     4801 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     4861 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     4921 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     4981 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     5041 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     5101 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     5161 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     5221 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     5281 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgccat
     5341 gcat
//
