LOCUS       sequence-245568-        6708 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-RPN13 (19-132) K83A in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6708)
  AUTHORS   Hemmis CW, Heard SC, Hill CP
  TITLE     Phosphorylation of Tyr950 in the proteasome scaffolding protein RPN2
            modulates its interaction with the ubiquitin receptor RPN13.
  JOURNAL   J Biol Chem. 2019 May 7. pii: AC119.008881. doi: 
            10.1074/jbc.AC119.008881.
  PUBMED    31064842
REFERENCE   2  (bases 1 to 6708)
  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: "J Biol 
            Chem. 2019 May 7. pii: AC119.008881. doi: 10.1074/jbc.AC119.008881."
COMMENT     Sequence Label: pET151-GST-PP-RPN13 (19-132) K83A
FEATURES             Location/Qualifiers
     source          1..6708
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     434..453
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(473..550)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     756..775
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     859..878
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        859..877
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    878..902
                     /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             917..939
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             968..1621
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             1628..1651
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(2059..2077)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2073..2120
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(2358..2376)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2444..2548
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2549..2617,2618..3409)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2767..2786)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3580..4168
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4069..4088
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4322..4339
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4354..4493
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4419..4438)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4579..4601
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4595..4786)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6098..6708,1..472))
                     /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"
ORIGIN
        1 tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag
       61 cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc
      121 gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca
      181 tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt
      241 gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc
      301 ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca
      361 ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct
      421 gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca
      481 ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt
      541 cgatggtgtc cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc
      601 agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg
      661 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc
      721 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca
      781 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag
      841 atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt
      901 cccctctaga aataattttg tttaacttta agaaggagat atacatatgg gaattgatcc
      961 cttcaccatg tcccctatac taggttattg gaaaattaag ggccttgtgc aacccactcg
     1021 acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc gcgatgaagg
     1081 tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc ttccttatta
     1141 tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata tagctgacaa
     1201 gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc ttgaaggagc
     1261 ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact ttgaaactct
     1321 caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag atcgtttatg
     1381 tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt tgtatgacgc
     1441 tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa aattagtttg
     1501 ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat ccagcaagta
     1561 tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc atcctccaaa
     1621 atcggatctg gaagttctgt tccaggggcc gtccaacaag tacttggtgg agtttcgggc
     1681 gggaaagatg tccctgaagg ggaccaccgt gactccggat aagcggaaag ggctggtgta
     1741 cattcagcag acggacgact cgcttattca cttctgctgg aaggacagga cgtccgggaa
     1801 cgtggaagac gacttgatca tcttccctga cgactgtgag ttcgcgcggg tgccgcagtg
     1861 ccccagcggg agggtctacg tgctgaagtt caaggcaggg tccaagcggc ttttcttctg
     1921 gatgcaggaa cccaagacag accaggatga ggagcattgc cggaaagtca acgagtatct
     1981 gaacaacccc ccgtaaaagg gcgagctcag atccggctgc taacaaagcc cgaaaggaag
     2041 ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     2101 gggtcttgag gggttttttg ctgaaaggag gaactatatc cggatatccc gcaagaggcc
     2161 cggcagtacc ggcataacca agcctatgcc tacagcatcc agggtgacgg tgccgaggat
     2221 gacgatgagc gcattgttag atttcataca cggtgcctga ctgcgttagc aatttaactg
     2281 tgataaacta ccgcattaaa gctagcttat cgatgataag ctgtcaaaca tgagaattaa
     2341 ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     2401 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     2461 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     2521 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     2581 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     2641 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     2701 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     2761 agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg
     2821 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     2881 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     2941 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     3001 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     3061 accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact
     3121 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     3181 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     3241 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     3301 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     3361 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     3421 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     3481 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     3541 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     3601 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     3661 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     3721 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     3781 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     3841 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     3901 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     3961 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     4021 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     4081 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     4141 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     4201 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     4261 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     4321 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     4381 tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca
     4441 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     4501 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     4561 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     4621 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
     4681 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     4741 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     4801 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     4861 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     4921 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     4981 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     5041 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     5101 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     5161 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     5221 tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgagatgc
     5281 gccgcgtgcg gctgctggag atggcggacg cgatggatat gttctgccaa gggttggttt
     5341 gcgcattcac agttctccgc aagaattgat tggctccaat tcttggagtg gtgaatccgt
     5401 tagcgaggtg ccgccggctt ccattcaggt cgaggtggcc cggctccatg caccgcgacg
     5461 caacgcgggg aggcagacaa ggtatagggc ggcgcctaca atccatgcca acccgttcca
     5521 tgtgctcgcc gaggcggcat aaatcgccgt gacgatcagc ggtccaatga tcgaagttag
     5581 gctggtaaga gccgcgagcg atccttgaag ctgtccctga tggtcgtcat ctacctgcct
     5641 ggacagcatg gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa gaatcataat
     5701 ggggaaggcc atccagcctc gcgtcgcgaa cgccagcaag acgtagccca gcgcgtcggc
     5761 cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac
     5821 gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt
     5881 cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc
     5941 tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc
     6001 ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc
     6061 ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     6121 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     6181 tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct
     6241 tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc
     6301 gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt
     6361 cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca
     6421 ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat
     6481 tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg
     6541 ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg
     6601 ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca
     6661 gatgctccac gcccagtcgc gtaccgtctt catgggagaa aataatac
//
