LOCUS       sequence-297107-        7260 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian overexpression vector for TRIM28; catalytically inactive 
            (mutated on C65A and C68A).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7260)
  AUTHORS   Rousseaux MW, de Haro M, Lasagna-Reeves CA, De Maio A, Park J, 
            Jafar-Nejad P, Al-Ramahi I, Sharma A, See L, Lu N, Vilanova-Velez L,
            Klisch TJ, Westbrook TF, Troncoso JC, Botas J, Zoghbi HY
  TITLE     TRIM28 regulates the nuclear accumulation and toxicity of both 
            alpha-synuclein and tau.
  JOURNAL   Elife. 2016 Oct 25;5. pii: e19809. doi: 10.7554/eLife.19809.
  PUBMED    27779468
REFERENCE   2  (bases 1 to 7260)
  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: "Elife."; 
            date: "2016-10-25"; volume: "5. pii: e19809. doi"; pages: " 
            10.7554/eLife.19809"
COMMENT     Sequence Label: pKH3-TRIM28-C65A/C68A
FEATURES             Location/Qualifiers
     source          1..7260
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     35..52
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    169..190
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(205..210,211..228,229..235)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    243..259
                     /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     248..270
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     267..283
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     267..283
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        312..691
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        692..895
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     846..866
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     892..916
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        1126..1144
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1126..1143
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     1157..1176
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             1237..1263
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1270..1296
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1303..1329
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      3858..3867
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3871..4005
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3908..3927)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3962..3981
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        4074..4431
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4074..4265
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4097..4117)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4282..4417
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4451..4468)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4451..4467)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4460..4482)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4680..5135
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4767..4786)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4977..4998
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5152..5171)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5271..5293
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5331..5349)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5417..5521
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5522..5590,5591..6382)
                     /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(5740..5759)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6553..7141
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7042..7061
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
       61 agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
      121 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
      181 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
      241 aattgtgagc ggataacaat ttcacacagg aaacagctat gacatgatta cgaattaatt
      301 cgagctcgcc cgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
      361 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
      421 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
      481 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
      541 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
      601 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
      661 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
      721 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
      781 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
      841 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctcgttt
      901 agtgaaccgt cagatcgcct ggagacgcca tccacgctgt tttgacctcc atagaagaca
      961 ccgggaccga tccagcctcc gcggccggga acggtgcatt ggaacgcgga ttccccgtgc
     1021 caagagtgac gtaagtaccg cctatagagt ctataggccc acccccttgg cttcgttaga
     1081 acgcggctac aattaataca taaccttatg tatcatacac atacgattta ggtgacacta
     1141 tagaataaca tccactttgc ctttctctcc acaggtgtcc actcccaggt ccaactgcac
     1201 ctcggttcta agcttctgca ggtcgactct agaatgtacc catacgatgt tccagattac
     1261 gctggatctt acccatacga tgttccagat tacgctggat cttacccata cgatgttcca
     1321 gattacgctg gatctatggc ggcctccgcg gcggcagcct cggcagcagc ggcctcggcc
     1381 gcctctggca gcccgggccc gggcgagggc tccgctggcg gcgaaaagcg ctccaccgcc
     1441 ccttcggccg cagcctcggc ctctgcctca gccgcggcgt cgtcgcccgc ggggggcggc
     1501 gccgaggcgc tggagctgct ggagcacgcc ggcgtggcca gagagcgcct gcgacccgag
     1561 agggagcccc gcctgctgcc ctgtttgcac tcggcctgta gtgcctgctt agggcccgcg
     1621 gcccccgccg ccgccaacag ctcgggggac ggcggggcgg cgggcgacgg caccgtggtg
     1681 gactgtcccg tgtgcaagca acagtgcttc tccaaagaca tcgtggagaa ttatttcatg
     1741 cgtgatagtg gcagcaaggc tgccaccgac gcccaggatg cgaaccagtg ctgcactagc
     1801 tgtgaggata atgccccagc caccagctac tgtgtggagt gctcggagcc tctgtgtgag
     1861 acctgtgtag aggcgcacca gcgggtgaag tacaccaagg accatactgt gcgctctact
     1921 gggccagcca agtctcggga tggtgaacgt actgtctatt gcaacgtaca caagcatgaa
     1981 ccccttgtgc tgttttgtga gagctgtgat actctcacct gccgagactg ccagctcaat
     2041 gcccacaagg accaccagta ccagttctta gaggatgcag tgaggaacca gcgcaagctc
     2101 ctggcctcac tggtgaagcg ccttggggac aaacatgcaa cattgcagaa gagcaccaag
     2161 gaggttcgca gctcaatccg ccaggtgtct gacgtacaga agcgtgtgca agtggatgtc
     2221 aagatggcca tcctgcagat catgaaggag ctgaataagc ggggccgtgt gctggtcaat
     2281 gatgcccaga aggtgactga ggggcagcag gagcgcctgg agcggcagca ctggaccatg
     2341 accaagatcc agaagcacca ggagcacatt ctgcgctttg cctcttgggc tctggagagt
     2401 gacaacaaca cagccctttt gctttctaag aagttgatct acttccagct gcaccgggcc
     2461 ctcaagatga ttgtggatcc cgtggagcca catggcgaga tgaaatttca gtgggacctc
     2521 aatgcctgga ccaagagtgc cgaggccttt ggcaagattg tggcagagcg tcctggcact
     2581 aactcaacag gccctgcacc catggcccct ccaagagccc cagggcccct gagcaagcag
     2641 ggctctggca gcagccagcc catggaggtg caggaaggct atggctttgg gtcaggagat
     2701 gatccctact caagtgcaga gccccatgtg tcaggtgtga aacggtcccg ctcaggtgag
     2761 ggcgaggtga gcggccttat gcgcaaggtg ccacgagtga gccttgaacg cctggacctg
     2821 gacctcacag ctgacagcca gccacccgtc ttcaaggtct tcccaggcag taccactgag
     2881 gactacaacc ttattgttat tgaacgtggc gctgccgctg cagctaccgg ccagccaggg
     2941 actgcgcctg caggaacccc tggtgcccca cccctggctg gcatggccat tgtcaaggag
     3001 gaggagacgg aggctgccat tggagcccct cctactgcca ctgagggccc tgagaccaaa
     3061 cctgtgctta tggctcttgc ggagggtcct ggtgctgagg gtccccgcct ggcctcacct
     3121 agtggcagca ccagctcagg gctggaggtg gtggctcctg agggtacctc agccccaggt
     3181 ggtggcccgg gaaccctgga tgacagtgcc accatttgcc gtgtctgcca gaagccaggc
     3241 gatctggtta tgtgcaacca gtgtgagttt tgtttccacc tggactgtca cctgccggcc
     3301 ctgcaggatg taccagggga ggagtggagc tgctcactct gccatgtgct ccctgacctg
     3361 aaggaggagg atggcagcct cagcctggat ggtgcagaca gcactggcgt ggtggccaag
     3421 ctctcaccag ccaaccagcg gaaatgtgag cgtgtactgc tggccctatt ctgtcacgaa
     3481 ccctgccgcc ccctgcatca gctggctacc gactccacct tctccctgga ccagcccggt
     3541 ggcaccctgg atctgaccct gatccgtgcc cgcctccagg agaagttgtc acctccctac
     3601 agctccccac aggagtttgc ccaggatgtg ggccgcatgt tcaagcaatt caacaagtta
     3661 actgaggaca aggcagacgt gcagtccatc atcggcctgc agcgcttctt cgagacgcgc
     3721 atgaacgagg ccttcggtga caccaagttc tctgctgtgc tggtggagcc cccgccgatg
     3781 agcctgcctg gtgctggcct gagttcccag gagctgtctg gtggccctgg tgatggcccc
     3841 tgagaattca atcgatggcc gccatggccc aacttgttta ttgcagctta taatggttac
     3901 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     3961 tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatcgggaa ttaattcggc
     4021 gcagcaccat ggcctgaaat aacctctgaa agaggaactt ggttaggtac cttctgaggc
     4081 ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     4141 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     4201 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     4261 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     4321 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     4381 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctgtta
     4441 acagcttggc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
     4501 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     4561 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc ctgatgcggt
     4621 attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta
     4681 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     4741 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     4801 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     4861 tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc
     4921 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     4981 actcttgttc caaactggaa caacactcaa ccctatctcg ggctattctt ttgatttata
     5041 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     5101 cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg
     5161 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     5221 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     5281 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat
     5341 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
     5401 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     5461 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     5521 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     5581 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     5641 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     5701 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     5761 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     5821 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     5881 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     5941 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     6001 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     6061 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     6121 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     6181 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     6241 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     6301 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     6361 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     6421 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     6481 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     6541 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     6601 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     6661 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     6721 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     6781 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     6841 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     6901 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     6961 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     7021 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     7081 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     7141 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     7201 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
//
