LOCUS       sequence-291199-        6174 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNot4_275-376.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6174)
  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 6174)
  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_275-376_AG
FEATURES             Location/Qualifiers
     source          1..6174
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      895..904
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1333..1359
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        complement(1730..1748)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1731..1748)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1768..1785)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1774..1998
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2044..2472
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2131..2150)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2341..2362
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2481..2501)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2486..2815
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2666..2801
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2728..2747
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2882..3676
                     /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(2936..2955)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3546..3565
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3850..3971
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3887..3906)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3941..3960
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4020..4036)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4020..4036)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4033..4055)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4044..4060
                     /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(4068..4074,4075..4092,4093..4098))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4113..4134
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4251..4268)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4422..5007)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4502..4521)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5178..5969,5970..6038))
                     /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     5801..5820
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6039..6143)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgccacc
      901 atggcttcta actttactca gttcgttctc gtcgacaatg gcggaactgg cgacgtgact
      961 gtcgccccaa gcaacttcgc taacggggtc gctgaatgga tcagctctaa ctcgcgatca
     1021 caggcttaca aagtaacctg tagcgttcgt cagagctctg cgcagaatcg caaatacacc
     1081 atcaaagtcg aggtgcctaa agtggcaacc cagactgttg gtggtgaaga gcttcctgta
     1141 gccggatgga gatcttactt aaatatggaa ctaaccattc caattttcgc cacgaattcc
     1201 gactgcgagc ttattgttaa ggcaatgcaa ggtctcctaa aagatggaaa cccgattccc
     1261 tcggccatcg cggccaactc cggcatctac ggaggtggag gtggatctgg tggatctcct
     1321 ccacatatgg cttacccata cgatgttcca gattacgctc ctccaaagct tggatccgaa
     1381 ttcccgggtc gactcgagcc cattgacaaa ccttcagatt ctctcagtat agggaacggt
     1441 gataattccc agcagatatc taacagtgat acgccttcac caccacctgg tttgtcaaaa
     1501 tccaatccag tcatccccat cagttcatcc aatcacagtg cacggtcccc ttttgaaggg
     1561 gcagtaacag agtcacagtc gttattctca gacaattttc gccatcccaa ccctatccca
     1621 agtgggcttc ctcctttccc cagctcccca cagacatcca gtgactggcc tacagcacca
     1681 gaaccacaga gcctcttcac atcataagcg gccgcctaga gggccctatt ctatagtgtc
     1741 acctaaatgc tagagctcgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg
     1801 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
     1861 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg
     1921 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg
     1981 atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc
     2041 cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     2101 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     2161 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     2221 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     2281 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     2341 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     2401 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     2461 ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     2521 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     2581 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     2641 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     2701 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc
     2761 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     2821 cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag gatcgtttcg
     2881 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
     2941 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
     3001 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
     3061 gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
     3121 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
     3181 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
     3241 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
     3301 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
     3361 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga
     3421 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
     3481 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
     3541 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
     3601 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
     3661 tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac
     3721 ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     3781 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     3841 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     3901 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     3961 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     4021 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     4081 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     4141 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4201 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     4261 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     4321 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     4381 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     4441 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     4501 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     4561 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     4621 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     4681 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     4741 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     4801 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     4861 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     4921 agctcttgat ccggcaaaca aaccaccgct ggtagcggtt tttttgtttg caagcagcag
     4981 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     5041 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5101 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     5161 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     5221 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     5281 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     5341 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     5401 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     5461 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     5521 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     5581 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     5641 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     5701 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     5761 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     5821 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     5881 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     5941 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     6001 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     6061 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     6121 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtc
//
