LOCUS       sequence-357460-        5729 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of HsNot1iso1_1356-1581.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5729)
  AUTHORS   Chen Y, Boland A, Kuzuoglu-Ozturk D, Bawankar P, Loh B, Chang CT, 
            Weichenrieder O, Izaurralde E
  TITLE     A DDX6-CNOT1 complex and W-binding pockets in CNOT9 reveal direct 
            links between miRNA target recognition and silencing.
  JOURNAL   Mol Cell. 2014 Jun 5;54(5):737-50. doi: 
            10.1016/j.molcel.2014.03.034. Epub 2014 Apr 24.
  PUBMED    24768540
REFERENCE   2  (bases 1 to 5729)
  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"; doi: 
            "10.1016/j.molcel.2014.03.034"; journalName: "Mol Cell"; date: 
            "2014-06-5- 5"; volume: "54"; issue: "5"; pages: "737-50"
COMMENT     Sequence Label: pnYC-NpM-HsNot1iso1_1356-1581_V
FEATURES             Location/Qualifiers
     source          1..5729
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1853..1872
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1961..1980
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1961..1979
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1980..2004
                     /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             2019..2041
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2048..3148
                     /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             3164..3187
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(3961..3979)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3975..4022
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(4197..4202,4203..4219,4220..4225))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     rep_origin      4337..4882
                     /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     4783..4802
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4999..5016
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        5408..5510
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             join(5511..5729,1..441)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(5577..5596)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 cgtatggcaa tgaaagacgg tgagctggtg atatgggata gtgttcaccc ttgttacacc
       61 gttttccatg agcaaactga aacgttttca tcgctctgga gtgaatacca cgacgatttc
      121 cggcagtttc tacacatata ttcgcaagat gtggcgtgtt acggtgaaaa cctggcctat
      181 ttccctaaag ggtttattga gaatatgttt ttcgtctcag ccaatccctg ggtgagtttc
      241 accagttttg atttaaacgt ggccaatatg gacaacttct tcgcccccgt tttcaccatg
      301 ggcaaatatt atacgcaagg cgacaaggtg ctgatgccgc tggcgattca ggttcatcat
      361 gccgtttgtg atggcttcca tgtcggcaga atgcttaatg aattacaaca gtactgcgat
      421 gagtggcagg gcggggcgta atttttttaa ggcagttatt ggtgccctta aacgcctggt
      481 tgctacgcct gaataagtga taataagcgg atgaatggca gaaattcgaa agcaaattcg
      541 acccggtcgt cggttcaggg cagggtcgtt aaatagccgc ttatgtctat tgctggttta
      601 ccggtttatt gactaccgga agcagtgtga ccgtgtgctt ctcaaatgcc tgaggccagt
      661 ttgctcaggc tctccccgtg gaggtaataa ttgacgatat gatcatttat tctgcctccc
      721 agagcctgat aaaaacggtt agcgcttcgt taatacagat gtaggtgttc cacagggtag
      781 ccagcagcat cctgcgatgc agatccggaa cataatggtg cagggcgctt gtttcggcgt
      841 gggtatggtg gcaggccccg tggccaggac ccaacgctgc ccgagatgcg ccgcgtgcgg
      901 ctgctggaga tggcggacgc gatggatatg ttctgccaag ggttggtttg cgcattcaca
      961 gttctccgca agaattgatt ggctccaatt cttggagtgg tgaatccgtt agcgaggtgc
     1021 cgccggcttc cattcaggtc gaggtggccc ggctccatgc accgcgacgc aacgcgggga
     1081 ggcagacaag gtatagggcg gcgcctacaa tccatgccaa cccgttccat gtgctcgccg
     1141 aggcggcata aatcgccgtg acgatcagcg gtccaatgat cgaagttagg ctggtaagag
     1201 ccgcgagcga tccttgaagc tgtccctgat ggtcgtcatc tacctgcctg gacagcatgg
     1261 cctgcaacgc gggcatcccg atgccgccgg aagcgagaag aatcataatg gggaaggcca
     1321 tccagcctcg cgtcgcgaac gccagcaaga cgtagcccag cgcgtcggcc gccatgccgg
     1381 cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag
     1441 cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc
     1501 gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca
     1561 tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg
     1621 agctgactgg gttgaaggct ctcaagggca tcggtcgacg ctctccctta tgcgactcct
     1681 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc gcaaggaatg
     1741 gtgcatgcaa ggagatggcg cccaacagtc ccccggccac ggggcctgcc accataccca
     1801 cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt
     1861 cggcgatata ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc
     1921 cggcgtagag gatcagacta agatctcgat cccgcgaaat taatacgact cactataggg
     1981 gaattgtgag cggataacaa ttccccacta gtaataattt tgtttaactt taagaaggag
     2041 atataccatg gaaactgaag aaggtaaact ggtaatctgg attaacggcg ataaaggcta
     2101 taacggtctc gctgaagtcg gtaagaaatt cgagaaagat accggaatta aagtcaccgt
     2161 tgagcatccg gataaactgg aagagaaatt cccacaggtt gcggcaactg gcgatggccc
     2221 tgacattatc ttctgggcac acgaccgctt tggtggctac gctcaatctg gcctgttggc
     2281 tgaaatcacc ccggacaaag cgttccagga caagctgtat ccgtttacct gggatgccgt
     2341 acgttacaac ggcaagctga ttgcttaccc gatcgctgtt gaagcgttat cgctgattta
     2401 taacaaagac ctgctgccga acccgccaaa aacctgggaa gagatcccgg cgctggataa
     2461 agaactgaaa gcgaaaggta agagcgccct gatgttcaac ctgcaagaac cgtacttcac
     2521 ctggccgctg attgctgctg acgggggtta tgcgttcaag tatgaaaacg gcaagtacga
     2581 cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg ggtctgacct tcctggttga
     2641 cctgattaaa aacaaacaca tgaatgcaga caccgattac tccatcgcag aagctgcctt
     2701 taataaaggc gaaacagcga tgaccatcaa cggcccgtgg gcatggtcca acatcgacac
     2761 cagcaaagtg aattatggtg taacggtact gccgaccttc aagggtcaac catccaaacc
     2821 gttcgttggc gtgctgagcg caggtattaa cgccgccagt ccgaacaaag agctggcgaa
     2881 agagttcctc gaaaactatc tgctgactga tgaaggtctg gaagcggtta ataaagacaa
     2941 accgctgggt gccgtagcgc tgaagtctta cgaggaagag ttggcgaaag atccacgtat
     3001 tgccgccacg atggaaaacg cccagaaagg tgaaatcatg ccgaacatcc cgcagatgtc
     3061 cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc gccagcggtc gtcagactgt
     3121 cgatgaagcc cttaaagacg cgcagactgg taccggcagc ggcctggaag ttctgttcca
     3181 ggggccccat atgctcgagc agtacagcta ccacgacatc aatgtctatt cccttgcggg
     3241 cttggcacca cacattactc taaatccaac aattcccttg tttcaggccc atccacagtt
     3301 gaagcagtgt gtgcgtcagg caattgaacg ggctgtccag gagctggtcc atcctgtggt
     3361 ggatcgatca attaagattg ccatgactac ttgtgagcaa atagtcagga aggattttgc
     3421 cctggattcg gaggaatctc gaatgcgaat agcagctcat cacatgatgc gtaacttgac
     3481 agctggaatg gctatgatta catgcaggga acctttgctc atgagcatat ctaccaactt
     3541 aaaaaacagt tttgcctcag cccttcgtac tgcttcccca caacaaagag aaatgatgga
     3601 tcaggcagct gctcaattag ctcaggacaa ttgtgagttg gcttgctgtt ttattcagaa
     3661 gactgcagta gaaaaagcag gccctgagat ggacaagaga ttagcaactg aatttgagct
     3721 gagaaaacat gctaggcaag aaggacgcag atactgtgat cctgttgttt taacatatca
     3781 agctgaacgg atgccagagc aaatcaggct gaaagttggt ggtgtggacc caaagcagtt
     3841 ggctgtttac gaagagtttg cacgcaatgt tcctggctaa ggatcctaat agtctagagc
     3901 tagccctagg agatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca
     3961 ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     4021 tgctgaaagg aggaactata tccggatatc ccgcaagagg cccggcagta ccggcataac
     4081 caagcctatg cctacagcat ccagggtgac ggtgccgagg atgacgatga gcgcattgtt
     4141 agatttcata cacggtgcct gactgcgtta gcaatttaac tgtgataaac taccgcatta
     4201 aagcttatcg atgataagct gtcaaacatg agaattacaa cttatatcgt atggggctga
     4261 cttcaggtgc tacatttgaa gagataaatt gcactgaaat caagaaatat tttatctgat
     4321 taataagatg atcttcttga gatcgttttg gtctgcgcgt aatctcttgc tctgaaaacg
     4381 aaaaaaccgc cttgcagggc ggtttttcga aggttctctg agctaccaac tctttgaacc
     4441 gaggtaactg gcttggagga gcgcagtcac caaaacttgt cctttcagtt tagccttaac
     4501 cggcgcatga cttcaagact aactcctcta aatcaattac cagtggctgc tgccagtggt
     4561 gcttttgcat gtctttccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     4621 tcggactgaa cggggggttc gtgcatacag tccagcttgg agcgaactgc ctacccggaa
     4681 ctgagtgtca ggcgtggaat gagacaaacg cggccataac agcggaatga caccggtaaa
     4741 ccgaaaggca ggaacaggag agcgcacgag ggagccgcca gggggaaacg cctggtatct
     4801 ttatagtcct gtcgggtttc gccaccactg atttgagcgt cagatttcgt gatgcttgtc
     4861 aggggggcgg agcctatgga aaaacggctt tgccgcggcc ctctcacttc cctgttaagt
     4921 atcttcctgg catcttccag gaaatctccg ccccgttcgt aagccatttc cgctcgccgc
     4981 agtcgaacga ccgagcgtag cgagtcagtg agcgaggaag cggaatatat cctgtatcac
     5041 atattctgct gacgcaccgg tgcagccttt tttctcctgc cacatgaagc acttcactga
     5101 caccctcatc agtgccaaca tagtaagcca gtatacactc ccctagcgct gatgtccggc
     5161 ggtgcttttg ccgttacgca ccaccccgtc agtagctgaa caggagggac agctgataga
     5221 aacagaagcc actggagcac ctcaaaaaca ccatcataca ctaaatcagt aagttggcag
     5281 catcacccga cgcactttgc gccgaataaa tacctgtgac ggaagatcac ttcgcagaat
     5341 aaataaatcc tggtgtccct gttgataccg ggaagccctg ggccaacttt tggcgaaaat
     5401 gagacgttga tcggcacgta agaggttcca actttcacca taatgaaata agatcactac
     5461 cgggcgtatt ttttgagtta tcgagatttt caggagctaa ggaagctaaa atggagaaaa
     5521 aaatcactgg atataccacc gttgatatat cccaatggca tcgtaaagaa cattttgagg
     5581 catttcagtc agttgctcaa tgtacctata accagaccgt tcagctggat attacggcct
     5641 ttttaaagac cgtaaagaaa aataagcaca agttttatcc ggcctttatt cacattcttg
     5701 cccgcctgat gaatgctcat ccggaattc
//
