LOCUS       sequence-316924-        7596 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNot4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7596)
  AUTHORS   Keskeny C, Raisch T, Sgromo A, Igreja C, Bhandari D, Weichenrieder 
            O, Izaurralde E
  TITLE     A conserved CAF40-binding motif in metazoan NOT4 mediates 
            association with the CCR4-NOT complex.
  JOURNAL   Genes Dev. 2019 Feb 1;33(3-4):236-252. doi: 10.1101/gad.320952.118. 
            Epub 2019 Jan 28.
  PUBMED    30692204
REFERENCE   2  (bases 1 to 7596)
  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."; date: "2019-02-1"; volume: "33(3-4):236-252. doi"; pages: " 
            10.1101/gad.320952.118. Epub 2019 Jan 28"
COMMENT     Sequence Label: pcDNA3.1-MS2-HA-HsNot4_AA
FEATURES             Location/Qualifiers
     source          1..7596
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      661..670
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1099..1125
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        complement(2915..2933)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2916..2933)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2953..2970)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2959..3183
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3229..3657
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3316..3335)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3526..3547
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3666..3686)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3671..4000
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3851..3986
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3913..3932
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4067..4861
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4121..4140)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4731..4750
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5035..5156
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5072..5091)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5126..5145
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5205..5221)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5205..5221)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5218..5240)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5229..5245
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5253..5259,5260..5277,5278..5283))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5298..5319
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5436..5453)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5607..6195)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5687..5706)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6366..7157,7158..7226))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6989..7008
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7227..7331)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7406..7425)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 gccaccatgg cttctaactt tactcagttc gttctcgtcg acaatggcgg aactggcgac
      721 gtgactgtcg ccccaagcaa cttcgctaac ggggtcgctg aatggatcag ctctaactcg
      781 cgatcacagg cttacaaagt aacctgtagc gttcgtcaga gctctgcgca gaatcgcaaa
      841 tacaccatca aagtcgaggt gcctaaagtg gcaacccaga ctgttggtgg tgaagagctt
      901 cctgtagccg gatggagatc ttacttaaat atggaactaa ccattccaat tttcgccacg
      961 aattccgact gcgagcttat tgttaaggca atgcaaggtc tcctaaaaga tggaaacccg
     1021 attccctcgg ccatcgcggc caactccggc atctacggag gtggaggtgg atctggtgga
     1081 tctcctccac atatggctta cccatacgat gttccagatt acgctcctcc aaagcttgga
     1141 tccgaattcc cgggtcgact cgagatgtct cgcagtcctg atgcgaagga agaccctgtg
     1201 gagtgccctc tttgcatgga gcccttggag atagatgata tcaacttttt cccttgcacc
     1261 tgtggctacc agatttgccg attttgttgg catcgaattc gcactgatga aaatgggctt
     1321 tgtcctgcat gtagaaagcc atatccagaa gacccagcag tttataaacc actctcccag
     1381 gaagagctgc aaaggataaa gaatgagaaa aaacagaaac aaaatgagag aaaacagaaa
     1441 atatcagaaa atcgcaaaca tttggctagt gtacgtgtcg tacaaaaaaa cctcgtcttt
     1501 gttgtaggtt tatctcagcg cctagcagac ccagaggttt taaaacgacc agaatatttt
     1561 gggaagtttg gtaaaataca taaagttgtc atcaataata gcacatcata tgcaggctca
     1621 cagggtccaa gtgccagtgc ttatgtaacc tatatccggt cagaagacgc tctcagagcc
     1681 atacagtgtg tcaacaatgt ggtagtagat ggcagaacac ttaaggcatc tctaggtaca
     1741 acaaaatact gcagttactt cttaaagaat atgcagtgtc caaaacctga ctgcatgtat
     1801 cttcatgaat tgggggatga ggcggccagc ttcacaaaag aggaaatgca ggcgggtaaa
     1861 caccaagaat atgaacagaa gctacttcaa gaattatata aattaaatcc caattttctt
     1921 cagctatcta cgggttcagt tgataaaaat aagaacaaag tgacaccact gcagaggtac
     1981 gataccccca ttgacaaacc ttcagattct ctcagtatag ggaacggtga taattcccag
     2041 cagatatcta acagtgatac gccttcacca ccacctggtt tgtcaaaatc caatccagtc
     2101 atccccatca gttcatccaa tcacagtgca cggtcccctt ttgaaggggc agtaacagag
     2161 tcacagtcgt tattctcaga caattttcgc catcccaacc ctatcccaag tgggcttcct
     2221 cctttcccca gctccccaca gacatccagt gactggccta cagcaccaga accacagagc
     2281 ctcttcacat cagaaacaat cccagtatca tcctctacag actggcaagc agcttttggc
     2341 tttggttctt ctaaacaacc agaggatgac ttgggttttg atcccttcga tgtcactcga
     2401 aaagccttag cagacctgat tgagaaggaa ctgtccgttc aagaccaacc ttccctttcg
     2461 cccacatctc ttcagaactc ctcttcacac actacaaccg ccaaaggtcc aggctctgga
     2521 ttcctgcatc ctgctgcagc tacaaatgcc aattctctca atagtacctt ttcagtcttg
     2581 ccccagaggt tccctcaatt tcagcagcac cgagcggttt ataattcatt cagttttcca
     2641 ggccaggcag cccgctatcc ttggatggcc tttccacgca atagcatcat gcacttgaac
     2701 cacacagcaa accccacctc aaatagtaat ttcttggact tgaatctccc gccacagcac
     2761 aacacaggtc tgggagggat ccctgtagca ggggaagaag aggtgaaggt ttcgaccatg
     2821 ccactgtcaa cctcttccca ttcattacaa caaggacagc agcctacaag tctccacact
     2881 actgtggcct aagcggccgc ctagagggcc ctattctata gtgtcaccta aatgctagag
     2941 ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     3001 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     3061 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg
     3121 acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta
     3181 tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta
     3241 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     3301 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     3361 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     3421 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     3481 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     3541 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     3601 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat
     3661 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     3721 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     3781 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     3841 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     3901 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa
     3961 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat
     4021 atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga
     4081 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     4141 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     4201 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc
     4261 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     4321 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     4381 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     4441 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     4501 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     4561 cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt
     4621 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     4681 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     4741 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     4801 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     4861 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     4921 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     4981 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     5041 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     5101 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     5161 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     5221 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     5281 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     5341 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     5401 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5461 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5521 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5581 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5641 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5701 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     5761 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     5821 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5881 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5941 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     6001 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     6061 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     6121 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     6181 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6241 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6301 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     6361 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6421 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     6481 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     6541 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     6601 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     6661 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     6721 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     6781 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     6841 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     6901 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     6961 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     7021 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     7081 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     7141 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     7201 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     7261 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     7321 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc
     7381 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
     7441 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
     7501 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
     7561 cgctgcttcg cgatgtacgg gccagatata cgcgtt
//
