LOCUS       sequence-194477-        6591 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for expression of NHA-tagged human TNRC6C CED in mammalian 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6591)
  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 6591)
  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-133
FEATURES             Location/Qualifiers
     source          1..6591
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        100..291
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     248..268
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(408..863)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(545..566)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     757..776
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    1049..1170
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1086..1105)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1140..1159
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(2236..2262)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             complement(2242..2268)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2305..2370)
                     /codon_start=1
                     /product="N-terminal RNA-binding domain from the lambda 
                     bacteriophage antiterminator protein N (Baron-Benhamou et 
                     al., 2004)"
                     /label=lambda N peptide
                     /translation="MDAQTRRRERRAEKQAQWKAAN"
     regulatory      2367..2376
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(2380..2398)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2441..2460)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          2443..2575
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     complement(2715..2739)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2736..2947)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2765..2785)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2948..3327
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(3538..4126)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3618..3637)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4297..5088,5089..5157))
                     /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     4920..4939
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5158..5262)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5330..5348
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5386..5408)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5508..5527
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     polyA_signal    5568..5616
                     /note="synthetic polyadenylation signal"
     CDS             complement(5680..6474)
                     /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(5791..5810)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6401..6420
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(6525..6591,1..291))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6539..6591,1..83)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 gcctcggcct ctgcataaat aaaaaaaatt agtcagccat ggggcggaga atgggcggaa
       61 ctgggcggag ttaggggcgg gatgggcgga gttaggggcg ggactatggt tgctgactaa
      121 ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt tccacacctg
      181 gttgctgact aattgagatg catgctttgc atacttctgc ctgctgggga gcctggggac
      241 tttccacacc ctaactgaca cacattccac agctggttct ttccgcctca gaaggtacct
      301 aaccaagttc ctctttcaga ggttatttca ggccatggtg ctgcgcagat ccgcgtatgc
      361 ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggaaa ttgtaagcgt
      421 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
      481 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
      541 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
      601 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
      661 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
      721 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
      781 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
      841 taatgcgccg ctacagggcg cgtccattcg ccattcaggc tgcgcaactg ttgggaaggg
      901 cgatcggtgc gggcctcttc gctattacgc cagcccggat cgatccttat cggattttac
      961 cacatttgta gaggttttac ttgctttaaa aaacctccca catctccccc tgaacctgaa
     1021 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     1081 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     1141 tggtttgtcc aaactcatca atgtatctta tcatgtctgc tcgaagcatt aaccctcact
     1201 aaagggaagc ggccgcggct ctcctggtgc tgatgatggc agcgcctaca gggactcccc
     1261 gctgagcagg tccccaggca gcagggtggt tagcggcgtg gggctgccta tcaccctgct
     1321 gtcgtcggcg ctgggcgggc cccacaggct gctggagtac tggggcaccc cgccccacag
     1381 cagctcactg ctccccttgc cacccaggca cggggcgttc cagtggccag cgtcgctgcg
     1441 taccaggcca tgggagctgc ccgctgcgct gagccgcggc tggctggacg cgctgctgga
     1501 ctgccagctg gaagtgggtg gcagcgcctg gccttgggct aagaagcgat tcacttcttc
     1561 ttcaccagcg aactcggcca ggatggtagt gtttcccagg acgcacatgt gcagagactt
     1621 ctgggccttg gcagcctcct ccttggagct gtaccggacc acagcattgc cttgagtcag
     1681 attcaggtgg aatgtgataa gaggcccatg ttgcaaacac aatgtccgca gtgtagaacc
     1741 atcaatctgg ggagtgaggt ttcgaagaac gagccagctg ctggttcttc ctgaggtgtc
     1801 ggtgctccag gcagaacccg aggagtagga gctggtccag ccgagggggc tggcacccca
     1861 ggtggaggag ggcttgggat tggttaaccc tggaggtggc ctcgtgggtg cagtactgtt
     1921 tctgggcacc ttccatagtt catgagacag agaggcttgc gtgtgggagg tagggccaga
     1981 ggaccacgtc gatttgatgt ctgacagttt acctccactc ttgaggaggt agcggttgac
     2041 atcctggatg gtggtgttga tggtaggccc agtggggaca ctgccaggag tgacgtcagg
     2101 gtcattctca gggtcaatat tctgcagtcc tttccatgga actcccggat ggaattctgg
     2161 gggccagttg atgctagagc catttgagat tttatcactg tcagatttgg cacggaattc
     2221 gacgggatcc gcgtcgagtg aagcgtaatc tggaacatca tatgggtagg ccatggtgta
     2281 agcttgggca gcctcgagcg gtgggtttgc agctttccat tgagcttgtt tctcagcgcg
     2341 acgctcacgt cgtcgtgttt gtgcgtccat ggtggctagc ctatagtgag tcgtattaag
     2401 tactctagcc ttaagagctg taattgaact gggagtggac acctgtggag agaaaggcaa
     2461 agtggatgtc agtaagacca ataggtgcct atcagaaacg caagagtctt ctctgtctcg
     2521 acaagcccag tttctattgg tctccttaaa cctgtcttgt aaccttgata cttacctgcc
     2581 cagtgcctca cgaccaactt ctgcaaggct accttaagag agtcactgca gcttaagttc
     2641 gagactgttg tgtcagaagc actgactgcg ttagcaattt aactgtgata aactaccgca
     2701 ataaagcttc tagtgatctg acggttcact aaacgagctc tgcttatata gacctcccac
     2761 cgtacacgcc taccgcccat ttgcgtcaac ggggcgggcg atcgcagttg ttacgacatt
     2821 ttggaaagtc ccgttgattt tggtgccaaa acaaactccc attgacgtca atggggtgga
     2881 gacttggaaa tccccgtgag tcaaaccgct atccacgccc attggtgtac tgccaaaacc
     2941 gcatcaccat ggtaatagcg atgactaata cgtagatgta ctgccaagta ggaaagtccc
     3001 gtaaggtcat gtactgggca taatgccagg cgggccattt accgtcattg acgtcaatag
     3061 ggggcggact tggcatatga tacacttgat gtactgccaa gtgggcagtt taccgtaaat
     3121 actccaccca ttgacgtcaa tggaaagtcc ctattggcgt tactatggga acatacgtca
     3181 ttattgacgt caatgggcgg gggtcgttgg gcggtcagcc aggcgggcca tttaccgtaa
     3241 gttatgtaac gcggaactcc atatatgggc tatgaactaa tgaccccgta attgattact
     3301 attaataact agtcaataat caatgccaac atggcggtca tattggacat gagccaatat
     3361 aaatgtacat attatgatat agatacaacg tatgcaatgg ccaatagcca atattgattt
     3421 atgctatata accaatgaat aatatggcta atggccaata ttgaagatct gtcgagccat
     3481 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     3541 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     3601 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3661 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3721 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3781 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     3841 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     3901 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     3961 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4021 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4081 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4141 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4201 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     4261 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     4321 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     4381 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     4441 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     4501 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     4561 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     4621 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     4681 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     4741 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     4801 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     4861 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     4921 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     4981 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5041 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5101 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     5161 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     5221 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     5281 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     5341 cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     5401 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     5461 gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg catcagagca
     5521 gattgtactg agagtgcacc atacgcggat ccttatcgct atcgattcac acaaaaaacc
     5581 aacacacaga tgtaatgaaa ataaagatat tttattgcgg ccatcgtgat ggcaggttgg
     5641 gcgtcgcttg gtcggtcatt tcgaacccca gagtcccgct cagaagaact cgtcaagaag
     5701 gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg
     5761 gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg ctatgtcctg
     5821 atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc ggccattttc
     5881 caccatgata ttcggcaagc aggcatcgcc atgggtcacg acgagatcct cgccgtcggg
     5941 catgcgcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat gctcttcgtc
     6001 cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct cgatgcgatg
     6061 tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc gccgcattgc
     6121 atcagccatg atggatactt tctcggcagg agcaaggtga gatgacagga gatcctgccc
     6181 cggcacttcg cccaatagca gccagtccct tcccgcttca gtgacaacgt cgagcacagc
     6241 tgcgcaagga acgcccgtcg tggccagcca cgatagccgc gctgcctcgt cctgcagttc
     6301 attcagggca ccggacaggt cggtcttgac aaaaagaacc gggcgcccct gcgctgacag
     6361 ccggaacacg gcggcatcag agcagccgat tgtctgttgt gcccagtcat agccgaatag
     6421 cctctccacc caagcggccg gagaacctgc gtgcaatcca tcttgttcaa tcatggtggc
     6481 tctagcctta agttcgagac tgttgtgtca gaagaatcaa gctttttgca aaagcctagg
     6541 cctccaaaaa agcctcctca ctacttctgg aatagctcag aggccgaggc g
//
