LOCUS       sequence-476849-        7387 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression for USP21 (S93A/S113A/S115A).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7387)
  TITLE     Mechanism of USP21 autoinhibition and histone H2AK119 
            deubiquitination
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7387)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pSR82: pET-SUMO-USP21 (S93A/S113A/S115A)
FEATURES             Location/Qualifiers
     source          1..7387
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(284..1366)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     1328..1347
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1367..1444)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1650..1669
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1757..1776
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1757..1775
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1776..1800
                     /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)."
     RBS             1815..1837
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1851..1868
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3836..3859
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             3871..3888
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3941..3959)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3955..4002
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4039..4494
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4126..4145)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4336..4357
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4520..4624
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4625..4693,4694..5485)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4843..4862)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5656..6244
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6145..6164
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6398..6415
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6430..6569
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6495..6514)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6655..6677
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6671..6862)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 agtgacgaag gcttgagcga gggcgtgcaa gattccgaat accgcaagcg acaggccgat
       61 catcgtcgcg ctccagcgaa agcggtcctc gccgaaaatg acccagagcg ctgccggcac
      121 ctgtcctacg agttgcatga taaagaagac agtcataagt gcggcgacga tagtcatgcc
      181 ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg gtcgagatcc
      241 cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca
      301 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
      361 tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat
      421 tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca
      481 gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag ctgtcttcgg
      541 tatcgtcgta tcccactacc gagatgtccg caccaacgcg cagcccggac tcggtaatgg
      601 cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc
      661 cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc
      721 gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca
      781 gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg
      841 cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga
      901 tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca
      961 cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg
     1021 cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca
     1081 ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg
     1141 gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca
     1201 gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt
     1261 cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag
     1321 agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc accaccctga
     1381 attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg
     1441 tgtccgggat ctcgacgctc tcccttatgc gactcctgca ttaggaagca gcccagtagt
     1501 aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga gatggcgccc
     1561 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
     1621 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
     1681 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat cgagatcgat
     1741 ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga taacaattcc
     1801 cctctagaaa taattttgtt taactttaag aaggagatat acatatgggc catcatcatc
     1861 atcatcatcc gtcaggagtc aagactgaga acaacgatca tattaatttg aaggtggcgg
     1921 ggcaggatgg ttctgtggtg cagtttaaga ttaagaggca tacaccactt agtaaactaa
     1981 tgaaagccta ttgtgaacga cagggattgt caatgaggca gatcagattc cgatttgacg
     2041 ggcaaccaat caatgaaaca gacacacctg cacagttgga aatggaggat gaagatacaa
     2101 ttgatgtgtt ccaacagcag acgggaggaa tgccccaggc ctctgagcac cgcctgggcc
     2161 gtacccgaga gccacctgtt aatatccagc cccgagtggg atccaagcta ccatttgccc
     2221 ccagggcccg cagcaaggag cgcagaaacc caggctctgg gccaaacccc atgttacgac
     2281 ctctgcctcc ccggccaggt ctgcctgatg aacggctcaa gaaactggag ctgggacggg
     2341 gacggacctc aggccctcgt cccagaggcc cccttcgagc agatcatggg gttcccctgc
     2401 ctggcgcgcc acccccaaca gtggctttgc ctctcccatc tcggaccaac ttagcccgtt
     2461 ccaaggcggt ggcgagtggg gacttgcgtc caatggggat tgccttggga gggcaccgtg
     2521 gcaccggaga gcttggggct gcactgagcc gcttggccct ccggcctgag ccacccactt
     2581 tgagacgtag cacttctctc cgccgcctag ggggctttcc tggaccccct accctgttca
     2641 gcatacggac agagccccct gcttcccatg gctccttcca catgatatcc gcccggtcct
     2701 ctgagccttt ctactctgat gacaagatgg ctcatcacac actccttctg ggctctggtc
     2761 atgttggcct tcgaaacctg ggaaacacgt gcttcctgaa tgctgtgctg cagtgtctga
     2821 gcagcactcg acctcttcgg gacttctgtc tgagaaggga cttccggcaa gaggtgcctg
     2881 gaggaggccg agcccaagag ctcactgaag cctttgcaga tgtgattggt gccctctggc
     2941 accctgactc ctgcgaagct gtgaatccta ctcgattccg agctgtcttc cagaaatatg
     3001 ttccctcctt ctctggatac agccagcagg atgcccaaga gttcctgaag ctcctcatgg
     3061 agcggctaca ccttgaaatc aaccgccgag gccgccgggc tccaccgata cttgccaatg
     3121 gtccagttcc ctctccaccc cgccgaggag gggctctgct agaagaacct gagttaagtg
     3181 atgatgaccg agccaaccta atgtggaaac gttacctgga gcgagaggac agcaagattg
     3241 tggacctgtt tgtgggccag ttgaaaagtt gtctcaagtg ccaggcctgt gggtatcgct
     3301 ccacgacctt cgaggttttt tgtgacctgt ccctgcccat ccccaagaaa ggatttgctg
     3361 ggggcaaggt gtctctgcgg gattgtttca accttttcac taaggaagaa gagctagagt
     3421 cggagaatgc cccagtgtgt gaccgatgtc ggcagaaaac tcgaagtacc aaaaagttga
     3481 cagtacaaag attccctcga atcctcgtgc tccatctgaa tcgattttct gcctcccgag
     3541 gctccatcaa aaaaagttca gtaggtgtag actttccact gcagcgactg agcctagggg
     3601 actttgccag tgacaaagcc ggaagtcctg tataccagct gtatgccctt tgcaaccact
     3661 caggcagcgt ccactatggc cactacacag ccctgtgccg gtgccagact ggttggcatg
     3721 tctacaatga ctctcgtgtc tcccctgtca gtgaaaacca ggtggcatcc agcgagggct
     3781 acgtgctgtt ctaccaactg atgcaggagc caccccggtg cctgtgatcg aggcctggag
     3841 ccacccgcag ttcgaaaagg gttaatcgag caccaccacc accaccactg agatccggct
     3901 gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca
     3961 taaccccttg gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata
     4021 tccggattgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     4081 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     4141 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     4201 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     4261 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     4321 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     4381 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     4441 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg
     4501 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     4561 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     4621 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     4681 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     4741 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     4801 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     4861 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     4921 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     4981 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     5041 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     5101 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     5161 gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     5221 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     5281 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     5341 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     5401 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     5461 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     5521 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     5581 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     5641 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     5701 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     5761 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
     5821 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     5881 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     5941 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     6001 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     6061 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     6121 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     6181 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     6241 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     6301 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     6361 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     6421 ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcaa
     6481 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc
     6541 tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc
     6601 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     6661 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa
     6721 gctcatcagc gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct
     6781 cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg
     6841 cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg
     6901 taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg
     6961 cccggttact ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga
     7021 gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg
     7081 gtagccagca gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc
     7141 gcgtttccag actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg
     7201 cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct
     7261 aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc
     7321 gcacccgtgg ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg
     7381 cgggacc
//
