LOCUS       sequence-291220-        7573 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNot4_275-575-del400-428.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7573)
  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 7573)
  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: pCIneo-V5-SBP-MBP-HsNot4_275-575-del400-428_AG
FEATURES             Location/Qualifiers
     source          1..7573
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..729
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     680..700
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     726..750
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          890..1022
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1005..1024
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1067..1085
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1098..1107
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1107..1148
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             1170..1283
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="MDEKTTGWGGGHVVEGLAGELEQLRARLEHHPQGQREP"
     regulatory      2265..2274
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3277..3398
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3288..3307)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3342..3361
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      3584..4039
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3671..3690)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3881..3902
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4156..4513
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4156..4347
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4179..4199)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4364..4499
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4564..5358
                     /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(4618..4637)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5228..5247
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5422..5470
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(5511..5530)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5630..5652
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5690..5708)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5776..5880
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5881..5949,5950..6741)
                     /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     complement(6099..6118)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6912..7500
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7401..7420
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactg
      661 cgatcgcccg ccccgttgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
      721 agcagagctc gtttagtgaa ccgtcagatc actagaagct ttattgcggt agtttatcac
      781 agttaaattg ctaacgcagt cagtgcttct gacacaacag tctcgaactt aagctgcagt
      841 gactctctta aggtagcctt gcagaagttg gtcgtgaggc actgggcagg taagtatcaa
      901 ggttacaaga caggtttaag gagaccaata gaaactgggc ttgtcgagac agagaagact
      961 cttgcgtttc tgataggcac ctattggtct tactgacatc cactttgcct ttctctccac
     1021 aggtgtccac tcccagttca attacagctc ttaaggctag agtacttaat acgactcact
     1081 ataggctagc aattgcggcc accatgggta agcctatccc taaccctctc ctcggtctcg
     1141 attctacgcc cggaagcggt agcggtacca tggacgagaa gaccaccggc tgggggggcg
     1201 gccacgtggt ggagggcctg gccggcgagc tggagcagct gcgggccagg ctggagcacc
     1261 accctcaggg ccagcgggag ccctccggcg gctgcaagcc cctcgacatg aaaatcgaag
     1321 aaggtaaact ggtaatctgg attaacggcg ataaaggcta taacggtctc gctgaagtcg
     1381 gtaagaaatt cgagaaagat accggaatta aagtcaccgt tgagcatccg gataaactgg
     1441 aagagaaatt cccgcaggtt gcggcaactg gcgatggccc tgacattatc ttctgggcac
     1501 acgaccgctt tggtggctac gctcaatctg gcctgttggc tgaaatcacc ccggacaaag
     1561 cgttccagga caagctgtat ccgtttacct gggatgccgt acgttacaac ggcaagctga
     1621 ttgcttaccc gatcgctgtt gaagcgttat cgctgattta taacaaagat ctgctgccga
     1681 acccgccaaa aacctgggaa gagatcccgg cgctggataa agaactgaaa gcgaaaggta
     1741 agagcgcgct gatgttcaac ctgcaagaac cgtacttcac ctggccgctg attgctgctg
     1801 acgggggtta tgcgttcaag tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg
     1861 ataacgctgg cgcgaaagcg ggtctgacct tcctggttga cctgattaaa agcaaacaca
     1921 tgaatgcaga caccgattac tccatcgcag aagctgcctt taataaaggc gaaacagcga
     1981 tgaccatcaa cggcccgtgg gcatggtcca acatcgacac cagcaaagtg aattatggtg
     2041 taacggtact gccgaccttc aagggtcaac catccaaacc gttcgttggc gtgctgagcg
     2101 caggtattaa cgccgccagt ccgaacaaag agctggcgaa agagttcctc gaaaactatc
     2161 tgctgactga tgaaggtctg gaagcggtta ataaagacaa accgctgggt gccgtagcgc
     2221 tgaagtctta cgaggaagag ttggcgaaag atccacgtat tgccgccacc atggaaaacg
     2281 cccagaaagg tgaaatcatg ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc
     2341 gtactgcggt gatcaacgcc gccagcggtc gtcagactgt cgatgaagcc ctgaaagacc
     2401 ctccactcga gcccattgac aaaccttcag attctctcag tatagggaac ggtgataatt
     2461 cccagcagat atctaacagt gatacgcctt caccaccacc tggtttgtca aaatccaatc
     2521 cagtcatccc catcagttca tccaatcaca gtgcacggtc cccttttgaa ggggcagtaa
     2581 cagagtcaca gtcgttattc tcagacaatt ttcgccatcc caaccctatc ccaagtgggc
     2641 ttcctccttt ccccagctcc ccacagacat ccagtgactg gcctacagca ccagaaccac
     2701 agagcctctt cacatcagaa acaatcccag tatcatcctc tacagactgg caagcagctt
     2761 ttggctttgg ttcttctaaa caaccacctt ccctttcgcc cacatctctt cagaactcct
     2821 cttcacacac tacaaccgcc aaaggtccag gctctggatt cctgcatcct gctgcagcta
     2881 caaatgccaa ttctctcaat agtacctttt cagtcttgcc ccagaggttc cctcaatttc
     2941 agcagcaccg agcggtttat aattcattca gttttccagg ccaggcagcc cgctatcctt
     3001 ggatggcctt tccacgcaat agcatcatgc acttgaacca cacagcaaac cccacctcaa
     3061 atagtaattt cttggacttg aatctcccgc cacagcacaa cacaggtctg ggagggatcc
     3121 ctgtagcagg ggaagaagag gtgaaggttt cgaccatgcc actgtcaacc tcttcccatt
     3181 cattacaaca aggacagcag cctacaagtc tccacactac tgtggcctaa gcggccgctt
     3241 ccctttagtg agggttaatg cttcgagcag acatgataag atacattgat gagtttggac
     3301 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     3361 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     3421 ttatgtttca ggttcagggg gagatgtggg aggtttttta aagcaagtaa aacctctaca
     3481 aatgtggtaa aatccgataa ggatcgatcc gggctggcgt aatagcgaag aggcccgcac
     3541 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggacgcgcc ctgtagcggc
     3601 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     3661 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     3721 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     3781 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     3841 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     3901 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     3961 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     4021 atattaacgc ttacaatttc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     4081 cacaccgcat acgcggatct gcgcagcacc atggcctgaa ataacctctg aaagaggaac
     4141 ttggttaggt accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg
     4201 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     4261 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     4321 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     4381 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     4441 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     4501 aggcttttgc aaaaagcttg attcttctga cacaacagtc tcgaacttaa ggctagagcc
     4561 accatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     4621 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     4681 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     4741 ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     4801 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     4861 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     4921 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     4981 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     5041 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc
     5101 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     5161 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     5221 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     5281 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     5341 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     5401 acctgccatc acgatggccg caataaaata tctttatttt cattacatct gtgtgttggt
     5461 tttttgtgtg aatcgatagc gataaggatc cgcgtatggt gcactctcag tacaatctgc
     5521 tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga
     5581 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     5641 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata
     5701 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     5761 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     5821 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     5881 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     5941 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     6001 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     6061 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     6121 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     6181 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     6241 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     6301 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     6361 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     6421 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     6481 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     6541 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     6601 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     6661 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     6721 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     6781 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     6841 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     6901 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     6961 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     7021 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     7081 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     7141 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     7201 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     7261 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     7321 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     7381 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     7441 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     7501 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     7561 gctcgacaga tct
//
