LOCUS       sequence-357789-        8239 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNot4-del190-274.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8239)
  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 8239)
  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-del190-274_AG
FEATURES             Location/Qualifiers
     source          1..8239
                     /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    3943..4064
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3954..3973)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4008..4027
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      4250..4705
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4337..4356)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4547..4568
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4822..5179
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4822..5013
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4845..4865)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5030..5165
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5230..6024
                     /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(5284..5303)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5894..5913
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6088..6136
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(6177..6196)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6296..6318
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6356..6374)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6442..6546
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6547..6615,6616..7407)
                     /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(6765..6784)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7578..8166
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8067..8086
                     /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 gatgtctcgc agtcctgatg cgaaggaaga ccctgtggag tgccctcttt
     2461 gcatggagcc cttggagata gatgatatca actttttccc ttgcacctgt ggctaccaga
     2521 tttgccgatt ttgttggcat cgaattcgca ctgatgaaaa tgggctttgt cctgcatgta
     2581 gaaagccata tccagaagac ccagcagttt ataaaccact ctcccaggaa gagctgcaaa
     2641 ggataaagaa tgagaaaaaa cagaaacaaa atgagagaaa acagaaaata tcagaaaatc
     2701 gcaaacattt ggctagtgta cgtgtcgtac aaaaaaacct cgtctttgtt gtaggtttat
     2761 ctcagcgcct agcagaccca gaggttttaa aacgaccaga atattttggg aagtttggta
     2821 aaatacataa agttgtcatc aataatagca catcatatgc aggctcacag ggtccaagtg
     2881 ccagtgctta tgtaacctat atccggtcag aagacgctct cagagccata cagtgtgtca
     2941 acaatgtggt agtagatggc agaacactta aggcatctgg tagcagcggt cccattgaca
     3001 aaccttcaga ttctctcagt atagggaacg gtgataattc ccagcagata tctaacagtg
     3061 atacgccttc accaccacct ggtttgtcaa aatccaatcc agtcatcccc atcagttcat
     3121 ccaatcacag tgcacggtcc ccttttgaag gggcagtaac agagtcacag tcgttattct
     3181 cagacaattt tcgccatccc aaccctatcc caagtgggct tcctcctttc cccagctccc
     3241 cacagacatc cagtgactgg cctacagcac cagaaccaca gagcctcttc acatcagaaa
     3301 caatcccagt atcatcctct acagactggc aagcagcttt tggctttggt tcttctaaac
     3361 aaccagagga tgacttgggt tttgatccct tcgatgtcac tcgaaaagcc ttagcagacc
     3421 tgattgagaa ggaactgtcc gttcaagacc aaccttccct ttcgcccaca tctcttcaga
     3481 actcctcttc acacactaca accgccaaag gtccaggctc tggattcctg catcctgctg
     3541 cagctacaaa tgccaattct ctcaatagta ccttttcagt cttgccccag aggttccctc
     3601 aatttcagca gcaccgagcg gtttataatt cattcagttt tccaggccag gcagcccgct
     3661 atccttggat ggcctttcca cgcaatagca tcatgcactt gaaccacaca gcaaacccca
     3721 cctcaaatag taatttcttg gacttgaatc tcccgccaca gcacaacaca ggtctgggag
     3781 ggatccctgt agcaggggaa gaagaggtga aggtttcgac catgccactg tcaacctctt
     3841 cccattcatt acaacaagga cagcagccta caagtctcca cactactgtg gcctaagcgg
     3901 ccgcttccct ttagtgaggg ttaatgcttc gagcagacat gataagatac attgatgagt
     3961 ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg
     4021 ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca
     4081 ttcattttat gtttcaggtt cagggggaga tgtgggaggt tttttaaagc aagtaaaacc
     4141 tctacaaatg tggtaaaatc cgataaggat cgatccgggc tggcgtaata gcgaagaggc
     4201 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatgga cgcgccctgt
     4261 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     4321 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     4381 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     4441 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     4501 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     4561 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     4621 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     4681 aacaaaatat taacgcttac aatttcctga tgcggtattt tctccttacg catctgtgcg
     4741 gtatttcaca ccgcatacgc ggatctgcgc agcaccatgg cctgaaataa cctctgaaag
     4801 aggaacttgg ttaggtacct tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt
     4861 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4921 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     4981 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     5041 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     5101 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     5161 aggcctaggc ttttgcaaaa agcttgattc ttctgacaca acagtctcga acttaaggct
     5221 agagccacca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     5281 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     5341 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     5401 aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     5461 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     5521 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     5581 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     5641 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     5701 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc
     5761 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     5821 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     5881 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     5941 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     6001 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     6061 cgcccaacct gccatcacga tggccgcaat aaaatatctt tattttcatt acatctgtgt
     6121 gttggttttt tgtgtgaatc gatagcgata aggatccgcg tatggtgcac tctcagtaca
     6181 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     6241 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     6301 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
     6361 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     6421 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     6481 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     6541 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     6601 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     6661 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     6721 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     6781 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     6841 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     6901 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     6961 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     7021 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     7081 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     7141 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     7201 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     7261 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     7321 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     7381 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     7441 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     7501 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     7561 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     7621 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     7681 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     7741 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     7801 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     7861 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     7921 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     7981 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     8041 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     8101 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     8161 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     8221 cacatggctc gacagatct
//
