LOCUS       sequence-322044-        5283 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses RNF4 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5283)
  AUTHORS   Guo L, Giasson BI, Glavis-Bloom A, Brewer MD, Shorter J, Gitler AD, 
            Yang X
  TITLE     A Cellular System that Degrades Misfolded Proteins and Protects 
            against Neurodegeneration.
  JOURNAL   Mol Cell. 2014 Jul 3;55(1):15-30. doi: 10.1016/j.molcel.2014.04.030.
            Epub 2014 May 29.
  PUBMED    24882209
REFERENCE   2  (bases 1 to 5283)
  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.2014.04.030"; journalName: "Mol Cell"; date: 
            "2014-07-3- 3"; volume: "55"; issue: "1"; pages: "15-30"
COMMENT     Sequence Label: HA-RNF4/pRK5
FEATURES             Location/Qualifiers
     source          1..5283
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     98..115
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    232..253
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(268..273,274..291,292..298)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    306..322
                     /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     311..333
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     330..346
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     330..346
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        375..754
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        755..958
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     909..929
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     955..979
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        1189..1207
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1189..1206
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     1220..1239
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             1295..1321
                     /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="YPYDVPDYA"
     CDS             1295..1321
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1910..1933
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      1944..1953
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1957..2091
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1994..2013)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2048..2067
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2160..2517
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2160..2351
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2183..2203)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2368..2503
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2537..2554)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2537..2553)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2546..2568)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2766..3221
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2853..2872)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3063..3084
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3238..3257)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3357..3379
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3417..3435)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3503..3607
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3608..3676,3677..4468)
                     /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(3826..3845)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4639..5227
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5128..5147
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
       61 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
      121 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
      181 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
      241 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
      301 tggaattgtg agcggataac aatttcacac aggaaacagc tatgacatga ttacgaatta
      361 attcgagctc gcccgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc
      421 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
      481 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
      541 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
      601 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
      661 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
      721 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa
      781 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
      841 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc
      901 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctcg
      961 tttagtgaac cgtcagatcg cctggagacg ccatccacgc tgttttgacc tccatagaag
     1021 acaccgggac cgatccagcc tccgcggccg ggaacggtgc attggaacgc ggattccccg
     1081 tgccaagagt gacgtaagta ccgcctatag agtctatagg cccaccccct tggcttcgtt
     1141 agaacgcggc tacaattaat acataacctt atgtatcata cacatacgat ttaggtgaca
     1201 ctatagaata acatccactt tgcctttctc tccacaggtg tccactccca ggtccaactg
     1261 cacctcggtt ctatcgattg aattcgccat gggctacccc tacgacgtgc ccgactacgc
     1321 cagatccatg agtacaagaa agcgtcgtgg tggagcaata aattctagac aagctcagaa
     1381 gcgaactcgg gaagcaacct ccacccccga gatctccttg gaagcagaac ccatagaact
     1441 cgtggaaact gctggagatg aaattgtgga cctcacttgt gaatctttag agcctgtggt
     1501 ggttgatctg actcacaatg actctgttgt gattgttgac gaaagaagaa gaccaaggag
     1561 gaatgctagg aggctgcccc aggaccatgc tgacagctgt gtggtgagca gtgacgatga
     1621 ggagttgtcc agggacagag acgtatatgt gactacccat actcccagaa acgccaggga
     1681 tgagggcgct acaggcctca ggccctcagg tactgtcagt tgtcccatct gcatggacgg
     1741 atactcagag atcgtgcaga atggacgtct catcgtttcc acagaatgcg gccatgtctt
     1801 ctgtagccag tgcctccgtg attccctgaa gaatgctaat acttgcccaa cttgtaggaa
     1861 aaagatcaac cacaaacggt accaccccat ttatatagtc gactgactcg actacaagga
     1921 cgacgatgac aagtgaaagc ttggccgcca tggcccaact tgtttattgc agcttataat
     1981 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     2041 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctggat cgggaattaa
     2101 ttcggcgcag caccatggcc tgaaataacc tctgaaagag gaacttggtt aggtaccttc
     2161 tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     2221 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     2281 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2341 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     2401 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     2461 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     2521 ctgttaacag cttggcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
     2581 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
     2641 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgcctga
     2701 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc
     2761 atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     2821 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     2881 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     2941 atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag
     3001 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     3061 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcgggct attcttttga
     3121 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     3181 atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac
     3241 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     3301 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     3361 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     3421 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     3481 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     3541 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     3601 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     3661 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     3721 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     3781 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     3841 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     3901 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     3961 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     4021 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     4081 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     4141 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     4201 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     4261 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     4321 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     4381 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     4441 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     4501 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     4561 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     4621 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     4681 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     4741 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     4801 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     4861 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     4921 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     4981 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     5041 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     5101 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     5161 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     5221 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     5281 tca
//
