LOCUS       sequence-455221-        7453 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pRS305 3XFLAG-HRV3C-Rpn5-VTENKIF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7453)
  AUTHORS   Greene ER, Goodall EA, de la Pena AH, Matyskiela ME, Lander GC, 
            Martin A
  TITLE     Specific lid-base contacts in the 26s proteasome control the 
            conformational switching required for substrate degradation.
  JOURNAL   Elife. 2019 Nov 28;8:e49806. doi: 10.7554/eLife.49806.
  PUBMED    31778111
REFERENCE   2  (bases 1 to 7453)
  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: "2019-11-28"; volume: "8:e49806. doi"; pages: " 
            10.7554/eLife.49806"
COMMENT     Sequence Label: pAM201
FEATURES             Location/Qualifiers
     source          1..7453
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(705..728)
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(738..803)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     promoter        complement(1339..1357)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1378..1394)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1378..1394)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1391..1413)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1402..1418
                     /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)."
     promoter        complement(join(1426..1432,1433..1450,1451..1456))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1471..1492
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1609..1626)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1780..2368)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1860..1879)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2539..3330,3331..3399))
                     /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     3162..3181
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3400..3504)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3572..3590
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3628..3650)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3750..3769
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(4263..5357)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(5358..5765)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     rep_origin      complement(6066..6521)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6203..6224)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6415..6434
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6647..6669
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6661..6678
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6662..6678
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6685..6704
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6685..6703
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 tattctctct tgtgcaaact aatttttacc agaagattat aatattctag cttcaatgac
       61 tcgtattttg gatttttaaa agtttttttc agaatttttc ttgaaagcac cgtggcttgg
      121 gaataatcac cttttaatat actcaattcc atttgctcta atataaactg aattttctca
      181 gacatttcca tggagccata ggtctcaacc tgtaactcac acaagatgtc tgcagcttca
      241 tcaatcttac cctcttcttt cttaatttct accaaatctt tggtgaccct agctctttcc
      301 acttctacag ctgctgcggc cgctgcagcc accctgatag tttcaatgac actaattctg
      361 gtgtttaaat ccaaagattt cgagcttttc aaatattcca taaccttttg tatcatatac
      421 tgaattgaca atttcaactg accatgcttt tttgagagta gagtcaattg ctcatttagg
      481 tcgtcccact tattccttga tgctagcaga tctacaatct tggccaaaac ttctttcgag
      541 gaggccagat ctgaagcttg tctggttttc ttctccaaca ctaacagttg gtctaaagca
      601 gagttacaat cattttgagc gagcgaatcg atcttaggaa actcttcctt caaaatttgg
      661 ctataatcct tgtcagcctt aattggtgca tctcttgaca tacctggacc ttgaaataaa
      721 acttccaaac cgcttccttt atcatcatca tctttgtaat caatatcatg atctttgtaa
      781 tcgccatcat gatctttgta atccatcttt ctctacgttt actctagcaa caaaaatggt
      841 actgagccca gcagtatatt cctctactct cttaattcaa gattatctct gatgtagtgt
      901 cctgttatgg tttttcctta gattcccctt tgcattttgc caccgaaaag tcaaaaagat
      961 cacatacaag acgggtaata ttggggcgat gagccgtaca gaacgagtga aaaattttag
     1021 gatttattag gaataataat gataatttag agatgagtat ttagcgagaa gcatctgaag
     1081 gttgcactaa ttattactgt aaatagtaca gttgagaagc aaatacaagg gaagatcaaa
     1141 tacatggccg gttcacaact taagaattta aaggccgcct tgaaggcccg tggactcaca
     1201 ggtcaaacaa acgtcaaaag taagaataaa aaaaattcca aaagacaagc gaaggagtat
     1261 gatcgtgaag aaaaaaaaaa ggccattgct gaaatcagag aagagtctcg agggggggcc
     1321 cggtaccagc ttttgttccc tttagtgagg gttaatttcg agcttggcgt aatcatggtc
     1381 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     1441 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     1501 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     1561 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     1621 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     1681 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     1741 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     1801 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     1861 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     1921 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     1981 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     2041 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     2101 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     2161 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     2221 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     2281 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     2341 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     2401 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     2461 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     2521 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     2581 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     2641 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     2701 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     2761 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     2821 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     2881 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     2941 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     3001 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     3061 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     3121 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     3181 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     3241 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     3301 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     3361 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     3421 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     3481 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga
     3541 aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct
     3601 cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac
     3661 agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt
     3721 tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac tgagagtgca
     3781 ccatatcgac tacgtcgtta aggccgtttc tgacagagta aaattcttga gggaactttc
     3841 accattatgg gaaatggttc aagaaggtat tgacttaaac tccatcaaat ggtcaggtca
     3901 ttgagtgttt tttatttgtt gtattttttt ttttttagag aaaatcctcc aatatataaa
     3961 ttaggaatca tagtttcatg attttctgtt acacctaact ttttgtgtgg tgccctcctc
     4021 cttgtcaata ttaatgttaa agtgcaattc tttttcctta tcacgttgag ccattagtat
     4081 caatttgctt acctgtattc ctttacatcc tcctttttct ccttcttgat aaatgtatgt
     4141 agattgcgta tatagtttcg tctaccctat gaacatattc cattttgtaa tttcgtgtcg
     4201 tttctattat gaatttcatt tataaagttt atgtacaaat atcataaaaa aagagaatct
     4261 ttttaagcaa ggattttctt aacttcttcg gcgacagcat caccgacttc ggtggtactg
     4321 ttggaaccac ctaaatcacc agttctgata cctgcatcca aaaccttttt aactgcatct
     4381 tcaatggcct taccttcttc aggcaagttc aatgacaatt tcaacatcat tgcagcagac
     4441 aagatagtgg cgatagggtt gaccttattc tttggcaaat ctggagcaga accgtggcat
     4501 ggttcgtaca aaccaaatgc ggtgttcttg tctggcaaag aggccaagga cgcagatggc
     4561 aacaaaccca aggaacctgg gataacggag gcttcatcgg agatgatatc accaaacatg
     4621 ttgctggtga ttataatacc atttaggtgg gttgggttct taactaggat catggcggca
     4681 gaatcaatca attgatgttg aaccttcaat gtagggaatt cgttcttgat ggtttcctcc
     4741 acagtttttc tccataatct tgaagaggcc aaaacattag ctttatccaa ggaccaaata
     4801 ggcaatggtg gctcatgttg tagggccatg aaagcggcca ttcttgtgat tctttgcact
     4861 tctggaacgg tgtattgttc actatcccaa gcgacaccat caccatcgtc ttcctttctc
     4921 ttaccaaagt aaatacctcc cactaattct ctgacaacaa cgaagtcagt acctttagca
     4981 aattgtggct tgattggaga taagtctaaa agagagtcgg atgcaaagtt acatggtctt
     5041 aagttggcgt acaattgaag ttctttacgg atttttagta aaccttgttc aggtctaaca
     5101 ctaccggtac cccatttagg accacccaca gcacctaaca aaacggcatc agccttcttg
     5161 gaggcttcca gcgcctcatc tggaagtgga acacctgtag catcgatagc agcaccacca
     5221 attaaatgat tttcgaaatc gaacttgaca ttggaacgaa catcagaaat agctttaaga
     5281 accttaatgg cttcggctgt gatttcttga ccaacgtggt cacctggcaa aacgacgatc
     5341 ttcttagggg cagacattag aatggtatat ccttgaaata tatatatata tattgctgaa
     5401 atgtaaaagg taagaaaagt tagaaagtaa gacgattgct aaccacctat tggaaaaaac
     5461 aataggtcct taaataatat tgtcaacttc aagtattgtg atgcaagcat ttagtcatga
     5521 acgcttctct attctatatg aaaagccggt tccggcgctc tcacctttcc tttttctccc
     5581 aatttttcag ttgaaaaagg tatatgcgtc aggcgacctc tgaaattaac aaaaaatttc
     5641 cagtcatcga atttgattct gtgcgatagc gcccctgtgt gttctcgtta tgttgaggaa
     5701 aaaaataatg gttgctaaga gattcgaact cttgcatctt acgatacctg agtattccca
     5761 cagttaactg cggtcaagat atttcttgaa tcaggcgcct tagaccgctc ggccaaacaa
     5821 ccaattactt gttgagaaat agagtataat tatcctataa atataacgtt tttgaacaca
     5881 catgaacaag gaagtacagg acaattgatt ttgaagagaa tgtggatttt gatgtaattg
     5941 ttgggattcc atttttaata aggcaataat attaggtatg tagatatact agaagttctc
     6001 ctcgaccggt cgatatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     6061 tcaggaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     6121 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     6181 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     6241 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     6301 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     6361 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     6421 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     6481 caccacaccc gccgcgctta atgcgccgct acagggcgcg tccattcgcc attcaggctg
     6541 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa
     6601 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt
     6661 tgtaaaacga cggccagtga attgtaatac gactcactat agggcgaatt ggagctcata
     6721 attttatctc acgtacgtaa atacacattt cgattagtga cagatttaaa atcaaaaccg
     6781 gatctgagat aatccgacag ttactcgaaa atctcttcat ttagcttgca gaccgtgcat
     6841 gatttcctct tttgtaatta aatggcctat tgtttctata tgttctaata gttcgtcaac
     6901 attatgtgac cattcgttca ataattggct tgagtttttt ggtttttcaa aattcacgat
     6961 tttggctggg cgattaactt tagcgtatat gatgccctgg tttaccaaat cactgatgta
     7021 tgtttccgtc tggctctccg ttaggtccag caattcattc aatcttagta aagtaattct
     7081 ggaatagtat tcggaaatga ctcttaaatt gtgctcgatg accctttttt gtaaatcttc
     7141 ccagtgatgc ttattagctt ctccaccaaa tgccaaatca tcctcattta agacgggctc
     7201 ataggttttt tgaacaattg gccatctcat caactcattc gtagtaaaca attttactaa
     7261 agattcttgg ctttctaatt ttttcaggtt gttatcattc tggattttgt gaattaaatc
     7321 attttgtaaa ttgccgtaag gtgaaaggac taagaaatat acaatgtgcg ataaaacagg
     7381 tttccactta gcctcatctg atttaatggc gtctgtttga taaatttctt gcagatactg
     7441 cgcaacttct agg
//
