LOCUS       sequence-245569-        6708 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-RPN13 (19-132) K97D 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) K97D
FEATURES             Location/Qualifiers
     source          1..6708
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     554..573
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(593..670)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     876..895
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     979..998
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        979..997
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    998..1022
                     /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             1037..1059
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1088..1741
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             1748..1771
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(2179..2197)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2193..2240
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(2478..2496)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2564..2668
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2669..2737,2738..3529)
                     /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(2887..2906)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3700..4288
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4189..4208
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4442..4459
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4474..4613
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4539..4558)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4699..4721
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4715..4906)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6218..6708,1..592))
                     /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 gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac
       61 ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac
      121 tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag
      181 cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc
      241 gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca
      301 tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt
      361 gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc
      421 ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca
      481 ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct
      541 gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca
      601 ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt
      661 cgatggtgtc cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc
      721 agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg
      781 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc
      841 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca
      901 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag
      961 atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt
     1021 cccctctaga aataattttg tttaacttta agaaggagat atacatatgg gaattgatcc
     1081 cttcaccatg tcccctatac taggttattg gaaaattaag ggccttgtgc aacccactcg
     1141 acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc gcgatgaagg
     1201 tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc ttccttatta
     1261 tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata tagctgacaa
     1321 gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc ttgaaggagc
     1381 ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact ttgaaactct
     1441 caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag atcgtttatg
     1501 tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt tgtatgacgc
     1561 tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa aattagtttg
     1621 ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat ccagcaagta
     1681 tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc atcctccaaa
     1741 atcggatctg gaagttctgt tccaggggcc gtccaacaag tacttggtgg agtttcgggc
     1801 gggaaagatg tccctgaagg ggaccaccgt gactccggat aagcggaaag ggctggtgta
     1861 cattcagcag acggacgact cgcttattca cttctgctgg aaggacagga cgtccgggaa
     1921 cgtggaagac gacttgatca tcttccctga cgactgtgag ttcaagcggg tgccgcagtg
     1981 ccccagcggg agggtctacg tgctggattt caaggcaggg tccaagcggc ttttcttctg
     2041 gatgcaggaa cccaagacag accaggatga ggagcattgc cggaaagtca acgagtatct
     2101 gaacaacccc ccgtaaaagg gcgagctcag atctggctgc taacaaagcc cgaaaggaag
     2161 ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     2221 gggtcttgag gggttttttg ctgaaaggag gaactatatc cggatatccc gcaagaggcc
     2281 cggcagtacc ggcataacca agcctatgcc tacagcatcc agggtgacgg tgccgaggat
     2341 gacgatgagc gcattgttag atttcataca cggtgcctga ctgcgttagc aatttaactg
     2401 tgataaacta ccgcattaaa gctagcttat cgatgataag ctgtcaaaca tgagaattaa
     2461 ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     2521 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     2581 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     2641 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     2701 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     2761 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     2821 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     2881 agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg
     2941 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     3001 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     3061 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     3121 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     3181 accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact
     3241 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     3301 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     3361 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     3421 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     3481 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     3541 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     3601 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     3661 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     3721 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     3781 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     3841 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     3901 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     3961 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     4021 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     4081 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     4141 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     4201 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     4261 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     4321 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     4381 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     4441 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     4501 tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca
     4561 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     4621 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     4681 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     4741 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
     4801 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     4861 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     4921 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     4981 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     5041 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     5101 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     5161 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     5221 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     5281 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     5341 tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgagatgc
     5401 gccgcgtgcg gctgctggag atggcggacg cgatggatat gttctgccaa gggttggttt
     5461 gcgcattcac agttctccgc aagaattgat tggctccaat tcttggagtg gtgaatccgt
     5521 tagcgaggtg ccgccggctt ccattcaggt cgaggtggcc cggctccatg caccgcgacg
     5581 caacgcgggg aggcagacaa ggtatagggc ggcgcctaca atccatgcca acccgttcca
     5641 tgtgctcgcc gaggcggcat aaatcgccgt gacgatcagc ggtccaatga tcgaagttag
     5701 gctggtaaga gccgcgagcg atccttgaag ctgtccctga tggtcgtcat ctacctgcct
     5761 ggacagcatg gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa gaatcataat
     5821 ggggaaggcc atccagcctc gcgtcgcgaa cgccagcaag acgtagccca gcgcgtcggc
     5881 cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac
     5941 gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt
     6001 cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc
     6061 tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc
     6121 ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc
     6181 ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     6241 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     6301 tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct
     6361 tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc
     6421 gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt
     6481 cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca
     6541 ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat
     6601 tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg
     6661 ctatcggctg aatttgattg cgagtgagat atttatgcca gccagcca
//
