LOCUS       sequence-101054-        7762 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express N-terminally FLAG tagged human chimera USP13/5-2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7762)
  AUTHORS   Liu Y, Soetandyo N, Lee JG, Liu L, Xu Y, Clemons WM Jr, Ye Y
  TITLE     USP13 antagonizes gp78 to maintain functionality of a chaperone in 
            ER-associated degradation.
  JOURNAL   Elife. 2014;3:e01369. doi: 10.7554/eLife.01369. Epub 2014 Jan 14.
  PUBMED    24424410
REFERENCE   2  (bases 1 to 7762)
  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: "2014"; volume: "3:e01369. doi"; pages: " 10.7554/eLife.01369.
            Epub 2014 Jan 14"
COMMENT     Sequence Label: PRK5-FLAG-USP13/5-2
FEATURES             Location/Qualifiers
     source          1..7762
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      918..927
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             927..950
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    1743..1754
                     /label=ZF binding site
                     /bound_moiety="CCR5TC zinc-finger domain"
                     /note="target sequence for the CCR5TC zinc-finger domain 
                     from the CCR5-224 (+) zinc-finger nuclease (Perez et al., 
                     2008; Gross et al., 2013)"
     promoter        complement(3553..3571)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3591..3608)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3597..3821
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3867..4295
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4304..4324)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4309..4639
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4489..4625
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4659..4676)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4659..4675)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4668..4690)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4888..5343
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5360..5379)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5479..5501
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5539..5557)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5625..5730
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5731..5799,5800..6591)
                     /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(5949..5968)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6762..7350
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7251..7270
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7499..7516
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7633..7654
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7669..7674,7675..7692,7693..7699)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7707..7723
                     /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     7712..7734
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7731..7747
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7731..7747
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc taagcttgcc gccatggact acaaggacga cgatgacaag gggtcgacaa
      961 tggcggagct gagtgaggag gcgctgctgt cagtattacc gacgatccgg gtccctaagg
     1021 ctggagaccg ggtccacaaa gacgagtgcg ccttctcctt cgacacgccg gagtctgagg
     1081 ggggcctcta catctgtatg aacacgtttc tgggctttgg gaaacagtat gtggagagac
     1141 atttcaataa gaccggccag cgagtctact tgcacctccg gcggacccgg cgcccgaaag
     1201 aggaggaccc tgctacaggc actggagacc caccccggaa gaagcccacg cggctggcta
     1261 ttggtgttga aggcgggttt gaccttagcg aggagaagtt tgaattagac gaggatgtga
     1321 agattgtcat tttgccagat tacctggaga ttgcccggga tggactgggg ggactgcctg
     1381 acattgtcag agatcgggtg accagtgcag tggaggccct actgtcggcc gactcagcct
     1441 cccgcaagca ggaggtgcag gcatgggatg gggaagtacg gcaggtgtct aagcatgcct
     1501 tcagcctcaa gcagttggac aaccctgctc gaatccctcc ctgtggctgg aagtgctcca
     1561 agtgtgacat gagagagaac ctgtggctca acctgactga tggctccatc ctctgtgggc
     1621 gacgctactt cgatggcagt gggggcaaca accacgctgt ggagcactac cgagagacag
     1681 gctacccgtt agctgtcaag ctgggcacca tcacccctga tggagctgac gtgtactcat
     1741 atgatgagga tgacatggtc ctggacccca gcctggctga gcacctgtcc cacttcggca
     1801 tcgacatgct gaagatgcag aagacagaca agacgatgac tgagttggag atagacatga
     1861 accagcggat tggtgaatgg gagctgatcc aggagtcagg tgtgccactc aagcccctgt
     1921 ttgggcctgg ctacacaggc atccggaacc tgggtaacag ctgctacctc aactctgtgg
     1981 tccaggtgct cttcagcatc cctgacttcc agaggaagta tgtggataag ctggagaaga
     2041 tcttccagaa tgccccgacg gaccctaccc aggatttcag cacccaggtg gccaagctgg
     2101 gccatggcct tctctccggg gagtattcca agccagtacc ggagtcgggc gatggggagc
     2161 gggtgccaga acagaaggaa gttcaacaga acgggatctc tccgcgcatg tttaaggcct
     2221 ttgtaagcaa gagccacccg gaattctcct ctaacaggca gcaagatgcc caggaattct
     2281 tcttgcacct ggtgaatcta gtagagagga accgcatcgg ctcagaaaac ccaagcgatg
     2341 tttttcgttt tttggtggaa gaacgcattc agtgctgtca gacccggaaa gtccgctaca
     2401 cggagagggt ggattacctg atgcagttac ctgtggccat ggaggcggca accaacaagg
     2461 atgaactgat cgcttatgaa ctaacgagaa gggaagcaga agcaaacaga agaccccttc
     2521 ctgagttggt acgtgccaag ataccattta gtgcctgcct tcaggccttc tctgaaccag
     2581 aaaatgttga tgatttctgg agcagtgccc tacaagcaaa gtctgcgggt gtgaaaacat
     2641 ctcgctttgc ttcattccct gaatacttgg tagtgcagat aaagaagttc acttttggtc
     2701 ttgactgggt tcccaaaaaa tttgatgttt ctattgatat gccagaccta cttgatatca
     2761 accatctccg agccaggggg ttacagccag gagaggaaga acttccagac atcagccccc
     2821 ccatagtcat tcctgatgac tcaaaagatc gcctgatgaa ccaattgata gacccatcag
     2881 acatcgatga gtcatcagtg atgcagctgg ccgagatggg tttcccgctg gaagcatgtc
     2941 gcaaggctgt gtacttcact ggaaatatgg gcgccgaggt ggccttcaac tggatcattg
     3001 ttcacatgga agagccagat tttgctgagc cgctgaccat gcctggttat ggaggggcag
     3061 cttctgctgg agcctctgtt tttggtgctt ctggactgga taaccaacct ccagaggaaa
     3121 tcgtagctat catcacctcc atgggatttc agcgaaatca ggctattcag gcactacgag
     3181 caacgaataa taacctggaa agagcactgg attggatctt tagccaccct gagtttgaag
     3241 aagacagtga ttttgtgatt gagatggaga ataatgccaa tgcaaacatt atttctgagg
     3301 ccaagcccga aggacctaga gtcaaggatg gatctggaac atatgagcta tttgcattca
     3361 tcagtcacat gggaacatcc acaatgagtg gtcattacat ttgccatatc aaaaaggaag
     3421 gaagatgggt gatttacaat gaccacaaag tttgtgcctc agaaaggccc cctaaagacc
     3481 tgggctacat gtacttttac cgcaggatac caagctaagc ggccgctcga gcatgcatct
     3541 agagggccct attctatagt gtcacctaaa tgctagagct cgctgatcag cctcgactgt
     3601 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     3661 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     3721 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     3781 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     3841 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     3901 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     3961 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     4021 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     4081 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     4141 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     4201 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     4261 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     4321 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     4381 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     4441 atgcatctca attagtcagc aaccatagtc acgcccctaa ctccgcccat cccgccccta
     4501 actccgccca gttccgccca ttctccgccc catggctgac taattttttt ttatttatgc
     4561 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     4621 aggcctaggc ttttgcaaaa agctgttaac agcttggcac tggccgtcgt tttacaacgt
     4681 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     4741 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     4801 ctgaatggcg aatggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     4861 caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg
     4921 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     4981 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     5041 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     5101 atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     5161 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     5221 ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     5281 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa
     5341 ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac
     5401 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     5461 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     5521 aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     5581 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     5641 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     5701 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     5761 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     5821 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     5881 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     5941 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     6001 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     6061 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     6121 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     6181 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     6241 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     6301 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     6361 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     6421 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     6481 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     6541 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     6601 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     6661 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     6721 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     6781 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     6841 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     6901 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     6961 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     7021 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     7081 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     7141 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     7201 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     7261 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     7321 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     7381 ctggcctttt gctggccttt tgctcacatg tcttcctgcg tatccnngat tctgtggata
     7441 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     7501 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
     7561 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
     7621 gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat
     7681 gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
     7741 ctatgacatg attacgaatt aa
//
