LOCUS       sequence-291144-        5560 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of DmNot4_813-836opt-L828E.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5560)
  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 5560)
  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: pnYC-NvM-DmNot4_813-836opt-L828E_AF
FEATURES             Location/Qualifiers
     source          1..5560
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2293..2312
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2401..2420
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2401..2419
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2420..2444
                     /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)."
     RBS             2459..2481
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2488..3588
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="METEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     CDS             3604..3624
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(3792..3810)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3806..3853
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(4028..4033,4034..4050,4051..4056))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     rep_origin      4168..4713
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     4614..4633
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4830..4847
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        5239..5341
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(5408..5427)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 gtatggcaat gaaagacggt gagctggtga tatgggatag tgttcaccct tgttacaccg
       61 ttttccatga gcaaactgaa acgttttcat cgctctggag tgaataccac gacgatttcc
      121 ggcagtttct acacatatat tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt
      181 tccctaaagg gtttattgag aatatgtttt tcgtctcagc caatccctgg gtgagtttca
      241 ccagttttga tttaaacgtg gccaatatgg acaacttctt cgcccccgtt ttcaccatgg
      301 gcaaatatta tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatcatg
      361 ccgtctgtga tggcttccat gtcggcagaa tgcttaatga attacaacag tactgcgatg
      421 agtggcaggg cggggcgtaa tttttttaag gcagttattg gtgcccttaa acgcctggtg
      481 ctacgcctga ataagtgata ataagcggat gaatggcaga aattcgaaag caaattcgac
      541 ccggtcgtcg gttcagggca gggtcgttaa atagccgctt atgtctattg ctggtttacc
      601 ggtttattga ctaccggaag cagtgtgacc gtgtgcttct caaatgcctg aggccagttt
      661 gctcaggctc tccccgtgga ggtaataatt gacgatatga tcatttattc tgcctcccag
      721 agcctgataa aaacggttag cgcttcgtta atacagatgt aggtgttcca cagggtagcc
      781 agcagcatcc tgcgatgcag atccggaaca taatggtgca gggcgcttgt ttcggcgtgg
      841 gtatggtggc aggccccgtg gccgggggac tgttgggcgc tgccggcacc tgtcctacga
      901 gttgcatgat aaagaagaca gtcataagtg cggcgacgat agtcatgccc cgcgcccacc
      961 ggaaggagct accggacagc ggtgcggact gttgtaactc agaataagaa atgaggccgc
     1021 tcatggcgtt gactctcagt catagtatcg tggtatcacc ggttggttcc actctctgtt
     1081 gcgggcaact tcagcagcac gtaggggact tccgcgtttc cagactttac gaaacacgga
     1141 aaccgaagac cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc
     1201 acgttcgctc gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta
     1261 gccgggtcct caacgacagg agcacgatca tgcgcacccg tggccaggac ccaacgctgc
     1321 ccgagatgcg ccgcgtgcgg ctgctggaga tggcggacgc gatggatatg ttctgccaag
     1381 ggttggtttg cgcattcaca gttctccgca agaattgatt ggctccaatt cttggagtgg
     1441 tgaatccgtt agcgaggtgc cgccggcttc cattcaggtc gaggtggccc ggctccatgc
     1501 accgcgacgc aacgcgggga ggcagacaag gtatagggcg gcgcctacaa tccatgccaa
     1561 cccgttccat gtgctcgccg aggcggcata aatcgccgtg acgatcagcg gtccagtgat
     1621 cgaagttagg ctggtaagag ccgcgagcga tccttgaagc tgtccctgat ggtcgtcatc
     1681 tacctgcctg gacagcatgg cctgcaacgc gggcatcccg atgccgccgg aagcgagaag
     1741 aatcataatg gggaaggcca tccagcctcg cgtcgcgaac gccagcaaga cgtagcccag
     1801 cgcgtcggcc gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg
     1861 accagtgacg aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc
     1921 gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg
     1981 cacctgtcct acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat
     2041 gccccgcgcc caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgacg
     2101 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     2161 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggccac
     2221 ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg
     2281 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
     2341 gatgccggcc acgatgcgtc cggcgtagag gatcagacta agatctcgat cccgcgaaat
     2401 taatacgact cactataggg gaattgtgag cggataacaa ttccccacta gtaataattt
     2461 tgtttaactt taagaaggag atataccatg gaaactgaag aaggtaaact ggtaatctgg
     2521 attaacggcg ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat
     2581 accggaatta aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt
     2641 gcggcaactg gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac
     2701 gctcaatctg gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat
     2761 ccgtttacct gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt
     2821 gaagcgttat cgctgattta taacaaagac ctgctgccga acccgccaaa aacctgggaa
     2881 gagatcccgg cgctggataa agaactgaaa gcgaaaggta agagcgccct gatgttcaac
     2941 ctgcaagaac cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag
     3001 tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg
     3061 ggtctgacct tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac
     3121 tccatcgcag aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg
     3181 gcatggtcca acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc
     3241 aagggtcaac catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt
     3301 ccgaacaaag agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg
     3361 gaagcggtta ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag
     3421 ttggcgaaag atccacgtat tgccgccacg atggaaaacg cccagaaagg tgaaatcatg
     3481 ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc
     3541 gccagcggtc gtcagactgt cgatgaagcc cttaaagacg cgcagactgg taccggcagc
     3601 ggcgaaaacc tttacttcca gggccatatg ctcgaggatg atgatctggg cttcgatccg
     3661 ttcgtggaaa cccagaaagg cgaagcggaa ctgatggaaa acgaagtgta aggatcctaa
     3721 tagtctagag ctagccctag gagatccggc tgctaacaaa gcccgaaagg aagctgagtt
     3781 ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
     3841 gaggggtttt ttgctgaaag gaggaactat atccggatat cccgcaagag gcccggcagt
     3901 accggcataa ccaagcctat gcctacagca tccagggtga cggtgccgag gatgacgatg
     3961 agcgcattgt tagatttcat acacggtgcc tgactgcgtt agcaatttaa ctgtgataaa
     4021 ctaccgcatt aaagcttatc gatgataagc tgtcaaacat gagaattaca acttatatcg
     4081 tatggggctg acttcaggtg ctacatttga agagataaat tgcactgaaa tcaagaaata
     4141 ttttatctga ttaataagat gatcttcttg agatcgtttt ggtctgcgcg taatctcttg
     4201 ctctgaaaac gaaaaaaccg ccttgcaggg cggtttttcg aaggttctct gagctaccaa
     4261 ctctttgaac cgaggtaact ggcttggagg agcgcagtca ccaaaacttg tcctttcagt
     4321 ttagccttaa ccggcgcatg acttcaagac taactcctct aaatcaatta ccagtggctg
     4381 ctgccagtgg tgcttttgca tgtctttccg ggttggactc aagacgatag ttaccggata
     4441 aggcgcagcg gtcggactga acggggggtt cgtgcataca gtccagcttg gagcgaactg
     4501 cctacccgga actgagtgtc aggcgtggaa tgagacaaac gcggccataa cagcggaatg
     4561 acaccggtaa accgaaaggc aggaacagga gagcgcacga gggagccgcc agggggaaac
     4621 gcctggtatc tttatagtcc tgtcgggttt cgccaccact gatttgagcg tcagatttcg
     4681 tgatgcttgt caggggggcg gagcctatgg aaaaacggct ttgccgcggc cctctcactt
     4741 ccctgttaag tatcttcctg gcatcttcca ggaaatctcc gccccgttcg taagccattt
     4801 ccgctcgccg cagtcgaacg accgagcgta gcgagtcagt gagcgaggaa gcggaatata
     4861 tcctgtatca catattctgc tgacgcaccg gtgcagcctt ttttctcctg ccacatgaag
     4921 cacttcactg acaccctcat cagtgccaac atagtaagcc agtatacact cccctagcgc
     4981 tgatgtccgg cggtgctttt gccgttacgc accaccccgt cagtagctga acaggaggga
     5041 cagctgatag aaacagaagc cactggagca cctcaaaaac accatcatac actaaatcag
     5101 taagttggca gcatcacccg acgcactttg cgccgaataa atacctgtga cggaagatca
     5161 cttcgcagaa taaataaatc ctggtgtccc tgttgatacc gggaagccct gggccaactt
     5221 ttggcgaaaa tgagacgttg atcggcacgt aagaggttcc aactttcacc ataatgaaat
     5281 aagatcacta ccgggcgtat tttttgagtt atcgagattt tcaggagcta aggaagctaa
     5341 aatggagaaa aaaatcactg gatataccac cgttgatata tcccaatggc atcgtaaaga
     5401 acattttgag gcatttcagt cagttgctca atgtacctat aaccagaccg ttcagctgga
     5461 tattacggcc tttttaaaga ccgtaaagaa aaataagcac aagttttatc cggcctttat
     5521 tcacattctt gcccgcctga tgaatgctca tccggaattc
//
