LOCUS       sequence-194413-        6966 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for expression of GST-tagged human TNRC6C CED 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-466
FEATURES             Location/Qualifiers
     source          1..6966
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     3..20
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4..20
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(1650..1667)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(1674..2327)
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MAPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     complement(1694..1716)
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     promoter        complement(2379..3557)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2388..3326
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(2411..2431)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     promoter        complement(3644..3840)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        complement(3652..3862)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3658..3793
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3894..3910)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3894..3910)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3907..3929)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3918..3934
                     /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(3942..3948,3949..3966,3967..3972))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3987..4008
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4125..4142)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4296..4884)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4376..4395)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5055..5846,5847..5915))
                     /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     5678..5697
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5916..6020)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6088..6106
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6144..6166)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6266..6285
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(6302..6757)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6439..6460)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6651..6670
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     join(6955..6966,1..11)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 gttgtaaaac gacggccagt gaattctttt tttttttttt tttttttttt tttttttttt
       61 ttttttggag gcaaacactt tattatcctg aagttcagtt acaatgtgac agtggagggg
      121 acactggtga gtggggggtg aagaggtgga gggtgacagt ggagcttcct ttcacacaca
      181 ggcctgacat gattcctcct gtctgctccc tcccacatcc ccagtccccg tccagcaagg
      241 caaatgtcca ggcacctgcc tggccctaaa aagagtcaca gtcctgaccc tcgtgggggc
      301 agggcaggtg tcccgaggcc acccagaaaa acaggagaca cgtcggtgat gttcgggagt
      361 caaaccatgt cccaaaagtc ttaagaagaa aaacgagatt ccccagcaaa ttcgtgggac
      421 cccagggcca tgcctcccag ctctggccac ctctgcttgt ccaggggcct cacagccaag
      481 gggatgcagg ggccgctgtg cagtcggctg aggccagagt gaggggtgca aggacccaga
      541 tggggaacac tgctgtttaa atattaaaca gggatttctt gtctcccacg tgggagacct
      601 gatactctca agggtcccag gtggggaccc tcactctaga gtcgcggccg cggctctcct
      661 ggtgctgatg atggcagcgc ctacagggac tccccgctga gcaggtcccc aggcagcagg
      721 gtggttagcg gcgtggggct gcctatcacc ctgctgtcgt cggcgctggg cgggccccac
      781 aggctgctgg agtactgggg caccccgccc cacagcagct cactgctccc cttgccaccc
      841 aggcacgggg cgttccagtg gccagcgtcg ctgcgtacca ggccatggga gctgcccgct
      901 gcgctgagcc gcggctggct ggacgcgctg ctggactgcc agctggaagt gggtggcagc
      961 gcctggcctt gggctaagaa gcgattcact tcttcttcac cagcgaactc ggccaggatg
     1021 gtagtgtttc ccaggacgca catgtgcaga gacttctggg ccttggcagc ctcctccttg
     1081 gagctgtacc ggaccacagc attgccttga gtcagattca ggtggaatgt gataagaggc
     1141 ccatgttgca aacacaatgt ccgcagtgta gaaccatcaa tctggggagt gaggtttcga
     1201 agaacgagcc agctgctggt tcttcctgag gtgtcggtgc tccaggcaga acccgaggag
     1261 taggagctgg tccagccgag ggggctggca ccccaggtgg aggagggctt gggattggtt
     1321 aaccctggag gtggcctcgt gggtgcagta ctgtttctgg gcaccttcca tagttcatga
     1381 gacagagagg cttgcgtgtg ggaggtaggg ccagaggacc acgtcgattt gatgtctgac
     1441 agtttacctc cactcttgag gaggtagcgg ttgacatcct ggatggtggt gttgatggta
     1501 ggcccagtgg ggacactgcc aggagtgacg tcagggtcat tctcagggtc aatattctgc
     1561 agtcctttcc atggaactcc cggatggaat tctgggggcc agttgatgct agagccattt
     1621 gagattttat cactgtcaga tttggcacgg gatccacgcg gaaccagatc cgattttgga
     1681 ggatggtcgc caccaccaaa cgtggcttgc cagccctgca aaggccatgc tatatacttg
     1741 ctggatttca agtacttatc aatttgtggg atagcttcaa tacgtttttt aaaacaaact
     1801 aattttggga acgcatccag gcacattggg tccatgtata aaacaacatc aagagcgtca
     1861 tacaacatga agtcaggatg ggttacatga tcaccattta aatatgtttt atgacataaa
     1921 cgatcttcga acattttcag catttcaggt agcttgctaa gaaaatcaac tttgagagtt
     1981 tcaaagtctt tactatatgc aattctcgaa acaccgtatc taatatccaa aaccgctcct
     2041 tcaagcattg aaatctctgc acgctctttt ggacaaccac ccaacatgtt gtgcttgtca
     2101 gctatataac gtatgatggc catagactgt gttaatttaa catcaccatc aatataataa
     2161 ggaagattgg gaaactccaa acccaattca aactttttgt ttcgccattt atcaccttca
     2221 tcgcgctcat acaaatgctc ttcatatttt tcttcaagat attccaaaag aagtcgagtg
     2281 ggttgcacaa ggcccttaat tttccaataa cctagtatag gggccattgt agatccagca
     2341 caatggatct cgaggtcgag ggatctctag agaattcctc acgacacctg aaatggaaga
     2401 aaaaaacttt gaaccactgt ctgaggcttg agaatgaacc aagatccaaa ctcaaaaagg
     2461 gcaaattcca aggagaatta catcaagtgc caagctggcc taacttcagt ctccacccac
     2521 tcagtgtggg gaaactccat cgcataaaac ccctcccccc aacctaaaga cgacgtactc
     2581 caaaagctcg agaactaatc gaggtgcctg gacggcgccc ggtactccgt ggagtcacat
     2641 gaagcgacgg ctgaggacgg aaaggccctt ttcctttgtg tgggtgactc acccgcccgc
     2701 tctcccgagc gccgcgtcct ccattttgag ctccctgcag cagggccggg aagcggccat
     2761 ctttccgctc acgcaactgg tgccgaccgg gccagccttg ccgcccaggg cggggcgata
     2821 cacggcggcg cgaggccagg caccagagca ggccggccag cttgagacta cccccgtccg
     2881 attctcggtg gccgcgctcg caggccccgc ctcgccgaac atgtgcgctg ggacgcacgg
     2941 gccccgtcgc cgcccgcggc cccaaaaacc gaaataccag tgtgcagatc ttggcccgca
     3001 tttacaagac tatcttgcca gaaaaaaagc gtcgcagcag gtcatcaaaa attttaaatg
     3061 gctagagact tatcgaaagc agcgagacag gcgcgaaggt gccaccagat tcgcacgcgg
     3121 cggccccagc gcccaggcca ggcctcaact caagcacgag gcgaaggggc tccttaagcg
     3181 caaggcctcg aactctccca cccacttcca acccgaagct cgggatcaag aatcacgtac
     3241 tgcagccagg tggaagtaat tcaaggcacg caagggccat aacccgtaaa gaggccaggc
     3301 ccgcgggaac cacacacggc acttacctgt gttctggcgg caaacccgtt gcgaaaaaga
     3361 acgttcacgg cgactactgc acttatatac ggttctcccc caccctcggg aaaaaggcgg
     3421 agccagtaca cgacatcact ttcccagttt accccgcgcc accttctcta ggcaccggtt
     3481 caattgccga cccctccccc caacttctcg gggactgtgg gcgatgtgcg ctctgcccac
     3541 tgacgggcac cggagccaat tcccactcct ttcaagacct agaaggtcca ttagctgcaa
     3601 agattcctct ctgtttaaaa ctttatccat ctttgcaaag ctttttgcaa aagcctaggc
     3661 ctccaaaaaa gcctcctcac tacttctgga atagctcaga ggccgaggcg gcctcggcct
     3721 ctgcataaat aaaaaaaatt agtcagccat ggggcggaga atgggcggaa ctgggcggag
     3781 ttaggggcgg gatgggcgga gttaggggcg ggactatggt tgctgactaa ttgagatgca
     3841 tgctttgcat acttctgcct gctggggagg gaattaagct tggcgtaatc atggtcatag
     3901 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     3961 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     4021 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     4081 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4141 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4201 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4261 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4321 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     4381 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     4441 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     4501 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     4561 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4621 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     4681 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     4741 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     4801 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     4861 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     4921 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     4981 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     5041 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5101 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5161 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     5221 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5281 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     5341 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     5401 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     5461 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     5521 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     5581 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     5641 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     5701 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     5761 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     5821 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     5881 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     5941 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     6001 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     6061 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     6121 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     6181 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     6241 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     6301 aaaattgtaa acgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
     6361 ttttttaacc aatagaccga aatcggcaaa atcccttata aatcaaaaga atagcccgag
     6421 atagagttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc
     6481 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc
     6541 aaatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc
     6601 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa
     6661 gcgaaaggag cgggcgctaa ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc
     6721 acacccgccg cgcttaatgc gccgctacag ggcgcgtact atggttgctt tgacgtatgc
     6781 ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc cattcgccat
     6841 tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc
     6901 tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt
     6961 cacgac
//
