LOCUS       sequence-101058-        7783 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express N-terminally FLAG tagged human chimera USP13/5-4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7783)
  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 7783)
  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-USP13/5-4
FEATURES             Location/Qualifiers
     source          1..7783
                     /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(3574..3592)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3612..3629)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3618..3842
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3888..4316
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4325..4345)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4330..4660
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4510..4646
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4680..4697)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4680..4696)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4689..4711)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4909..5364
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5381..5400)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5500..5522
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5560..5578)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5646..5751
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5752..5820,5821..6612)
                     /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(5970..5989)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6783..7371
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7272..7291
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7520..7537
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7654..7675
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7690..7695,7696..7713,7714..7720)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7728..7744
                     /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     7733..7755
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7752..7768
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7752..7768
                     /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 gttcaagatg gcattgcccc tcggatgttc aaggccctca
     2221 tcggcaaggg ccaccctgaa ttctccacca accggcagca ggatgcccag gagttcttcc
     2281 ttcaccttat caacatggtg gagaggaatt gccggagctc tgaaaatcct aatgaagtgt
     2341 tccgcttctt ggtggaggaa aagatcaagt gcctggccac agagaaggtg aagtacaccc
     2401 agcgagttga ctacatcatg cagctgcctg tgcccatgga tgcagccctt aacaaagagg
     2461 agcttctgga gtacgaggag aagaagcggc aagccgaaga ggagaagatg gcactgccag
     2521 aactggttcg ggcccaggtg cccttcagct cttgcctgga ggcctacggg gcccctgagc
     2581 aggtcgatga cttctggagc acggccctgc aggccaagtc agtagctgtc aagaccacac
     2641 gatttgcctc attccctgac tacctggtca tccagatcaa gaagttcacc ttcggcttag
     2701 actgggtgcc caagaaactg gatgtgtcca tcgagatgcc agaggagctc gacatctccc
     2761 agttgagggg cacagggctg cagcccggag aggaggagct gccagacatt gccccacccc
     2821 tggtcactcc ggatgagccc aaaggtagcc ttggtttcta tggcaacgaa gacgaagact
     2881 ccttctgctc ccctcacttc tcctctccga catcgcccat gctggatgaa tcagtcatca
     2941 tccagctggt ggagatggga ttccctatgg acgcctgccg caaagctgtc tactacacgg
     3001 gcaacagcgg ggctgaggcc gccatgaact gggtcatgtc acacatggat gatccagatt
     3061 ttgcaaaccc cctcatcctg cctggctcta gtgggccggg ctccacaagc tctggactgg
     3121 ataaccaacc tccagaggaa atcgtagcta tcatcacctc catgggattt cagcgaaatc
     3181 aggctattca ggcactacga gcaacgaata ataacctgga aagagcactg gattggatct
     3241 ttagccaccc tgagtttgaa gaagacagtg attttgtgat tgagatggag aataatgcca
     3301 atgcaaacat tatttctgag gccaagcccg aaggacctag agtcaaggat ggatctggaa
     3361 catatgagct atttgcattc atcagtcaca tgggaacatc cacaatgagt ggtcattaca
     3421 tttgccatat caaaaaggaa ggaagatggg tgatttacaa tgaccacaaa gtttgtgcct
     3481 cagaaaggcc ccctaaagac ctgggctaca tgtactttta ccgcaggata ccaagctaag
     3541 cggccgctcg agcatgcatc tagagggccc tattctatag tgtcacctaa atgctagagc
     3601 tcgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     3661 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     3721 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     3781 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     3841 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     3901 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     3961 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     4021 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     4081 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     4141 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     4201 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     4261 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     4321 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     4381 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     4441 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cacgccccta
     4501 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     4561 ctaatttttt tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa
     4621 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctgttaa cagcttggca
     4681 ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc
     4741 cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc
     4801 ccttcccaac agttgcgcag cctgaatggc gaatggcgcc tgatgcggta ttttctcctt
     4861 acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac gcgccctgta
     4921 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     4981 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     5041 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     5101 acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca tcgccctgat
     5161 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     5221 aaactggaac aacactcaac cctatctcgg gctattcttt tgatttataa gggattttgc
     5281 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     5341 acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc tctgatgccg
     5401 catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
     5461 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     5521 ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt
     5581 tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa
     5641 atgtgcgcgg aacccctatt ttgtttattt ttctaaatac attcaaatat gtatccgctc
     5701 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     5761 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     5821 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     5881 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     5941 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     6001 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     6061 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     6121 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     6181 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     6241 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     6301 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     6361 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     6421 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     6481 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     6541 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     6601 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     6661 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     6721 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     6781 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     6841 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     6901 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     6961 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     7021 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     7081 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     7141 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     7201 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     7261 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     7321 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     7381 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gtcttcctgc
     7441 gtatccnnga ttctgtggat accgtattac cgcctttgag tgagctgata ccgctcgccg
     7501 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     7561 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     7621 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
     7681 accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
     7741 acaatttcac acaggaaaca gctatgacat gattacgaat taa
//
