LOCUS       sequence-194412-        6966 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for expression of GST-tagged human TNRC6C 7W7A CED with 
            seven mutated W-motifs in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6966)
  AUTHORS   Chekulaeva M, Mathys H, Zipprich JT, Attig J, Colic M, Parker R, 
            Filipowicz W
  TITLE     miRNA repression involves GW182-mediated recruitment of CCR4-NOT 
            through conserved W-containing motifs.
  JOURNAL   Nat Struct Mol Biol. 2011 Oct 7;18(11):1218-26. doi: 
            10.1038/nsmb.2166.
  PUBMED    21984184
REFERENCE   2  (bases 1 to 6966)
  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: "10.1038/nsmb"; 
            journalName: "Nat Struct Mol Biol"; date: "2011-10-7- 7"; volume: 
            "18"; issue: "11"; pages: "1218-26"
COMMENT     Sequence Label: pMCh-633
FEATURES             Location/Qualifiers
     source          1..6966
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(52..69)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(52..68)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(61..83)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      281..736
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(368..387)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     578..599
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(753..772)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     872..894
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(932..950)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1018..1122
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1123..1191,1192..1983)
                     /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(1341..1360)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2154..2742
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2643..2662
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2896..2913
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3030..3051
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3066..3071,3072..3089,3090..3096)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3104..3120
                     /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)."
     primer_bind     3109..3131
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3128..3144
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3128..3144
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        3176..3386
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        3198..3394
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3245..3380
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3481..4659
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          3712..4650
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     4607..4627
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             4711..5364
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MAPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     5322..5344
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             5371..5388
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
ORIGIN
        1 cctccaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaagaatt cactggccgt
       61 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
      121 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
      181 acagttgcgc agcctgaatg gcgaatggcg cctgatgcgg tattttctcc ttacgcatct
      241 gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca
      301 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta
      361 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
      421 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
      481 cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt
      541 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
      601 acaacactca actctatctc gggctattct tttgatttat aagggatttt gccgatttcg
      661 gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
      721 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
      781 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
      841 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
      901 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
      961 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     1021 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     1081 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     1141 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     1201 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     1261 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     1321 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     1381 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     1441 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     1501 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     1561 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     1621 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     1681 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     1741 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     1801 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     1861 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     1921 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     1981 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     2041 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     2101 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     2161 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     2221 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     2281 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     2341 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     2401 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     2461 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     2521 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     2581 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     2641 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     2701 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     2761 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     2821 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     2881 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     2941 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     3001 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     3061 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     3121 tttcacacag gaaacagcta tgaccatgat tacgccaagc ttaattccct ccccagcagg
     3181 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     3241 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     3301 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
     3361 aggaggcttt tttggaggcc taggcttttg caaaaagctt tgcaaagatg gataaagttt
     3421 taaacagaga ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt
     3481 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
     3541 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
     3601 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
     3661 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
     3721 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
     3781 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
     3841 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
     3901 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
     3961 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
     4021 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
     4081 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
     4141 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
     4201 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     4261 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     4321 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     4381 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     4441 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     4501 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     4561 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     4621 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattctcta gagatccctc
     4681 gacctcgaga tccattgtgc tggatctaca atggccccta tactaggtta ttggaaaatt
     4741 aagggccttg tgcaacccac tcgacttctt ttggaatatc ttgaagaaaa atatgaagag
     4801 catttgtatg agcgcgatga aggtgataaa tggcgaaaca aaaagtttga attgggtttg
     4861 gagtttccca atcttcctta ttatattgat ggtgatgtta aattaacaca gtctatggcc
     4921 atcatacgtt atatagctga caagcacaac atgttgggtg gttgtccaaa agagcgtgca
     4981 gagatttcaa tgcttgaagg agcggttttg gatattagat acggtgtttc gagaattgca
     5041 tatagtaaag actttgaaac tctcaaagtt gattttctta gcaagctacc tgaaatgctg
     5101 aaaatgttcg aagatcgttt atgtcataaa acatatttaa atggtgatca tgtaacccat
     5161 cctgacttca tgttgtatga cgctcttgat gttgttttat acatggaccc aatgtgcctg
     5221 gatgcgttcc caaaattagt ttgttttaaa aaacgtattg aagctatccc acaaattgat
     5281 aagtacttga aatccagcaa gtatatagca tggcctttgc agggctggca agccacgttt
     5341 ggtggtggcg accatcctcc aaaatcggat ctggttccgc gtggatcccg tgccaaatct
     5401 gacagtgata aaatctcaaa tggctctagc atcaactggc ccccagaatt ccatccggga
     5461 gttccatgga aaggactgca gaatattgac cctgagaatg accctgacgt cactcctggc
     5521 agtgtcccca ctgggcctac catcaacacc accatccagg atgtcaaccg ctacctcctc
     5581 aagagtggag gtaaactgtc agacatcaaa tcgacggcct cctctggccc tacctcccac
     5641 acgcaagcct ctctgtctca tgaactatgg aaggtgccca gaaacagtac tgcacccacg
     5701 aggccacctc cagggttaac caatcccaag ccctcctcca ccgccggtgc cagccccctc
     5761 ggcgccacca gctcctactc ctcgggttct gccgccagca ccgacacctc aggaagaacc
     5821 agcagctggc tcgttcttcg aaacctcact ccccagattg atggttctac actgcggaca
     5881 ttgtgtttgc aacatgggcc tcttatcaca ttccacctga atctgactca aggcaatgct
     5941 gtggtccggt acagctccaa ggaggaggct gccaaggccc agaagtctct gcacatgtgc
     6001 gtcctgggaa acactaccat cctggccgag ttcgctggtg aagaagaagt gaatcgcttc
     6061 ttagcccaag gccaggcgct gccacccact tccagcgccc agtccagcag cgcgtccagc
     6121 cagccgcggc tcagcgcagc gggcagctcc catggcctgg tacgcagcga cgctggccac
     6181 tggaacgccc cgtgcctggg tggcaagggg agcagtgagc tgctggccgg cggggtgccc
     6241 cagtactcca gcagcctggc cggcccgccc agcgccgacg acagcagggt gataggcagc
     6301 cccacgccgc taaccaccct gctgcctggg gacctgctca gcggggagtc cctgtaggcg
     6361 ctgccatcat cagcaccagg agagccgcgg ccgcgactct agagtgaggg tccccacctg
     6421 ggacccttga gagtatcagg tctcccacgt gggagacaag aaatccctgt ttaatattta
     6481 aacagcagtg ttccccatct gggtccttgc acccctcact ctggcctcag ccgactgcac
     6541 agcggcccct gcatcccctt ggctgtgagg cccctggaca agcagaggtg gccagagctg
     6601 ggaggcatgg ccctggggtc ccacgaattt gctggggaat ctcgtttttc ttcttaagac
     6661 ttttgggaca tggtttgact cccgaacatc accgacgtgt ctcctgtttt tctgggtggc
     6721 ctcgggacac ctgccctgcc cccacgaggg tcaggactgt gactcttttt agggccaggc
     6781 aggtgcctgg acatttgcct tgctggacgg ggactgggga tgtgggaggg agcagacagg
     6841 aggaatcatg tcaggcctgt gtgtgaaagg aagctccact gtcaccctcc acctcttcac
     6901 cccccactca ccagtgtccc ctccactgtc acattgtaac tgaacttcag gataataaag
     6961 tgtttg
//
