LOCUS       sequence-455219-        6736 bp    DNA     circular SYN 15-APR-2025
DEFINITION  pRS316 Rpn6 promoter-Rpn5 ORF-Rpn5 terminator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6736)
  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 6736)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pAM202
FEATURES             Location/Qualifiers
     source          1..6736
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      111..566
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(198..217)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     408..429
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(697..1500)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(1501..1716)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(1740..1759)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1859..1881
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1919..1937)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    1978..2481
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        2518..2622
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2623..2691,2692..3483)
                     /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(2841..2860)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3654..4242
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4143..4162
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4396..4413
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4530..4551
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4566..4571,4572..4589,4590..4596)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4604..4620
                     /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     4609..4631
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4628..4644
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4628..4644
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        4665..4683
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(6664..6683)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6665..6683)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6690..6707)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6690..6706)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6699..6721)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
       61 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg acgcgccctg
      121 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
      181 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
      241 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
      301 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
      361 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
      421 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
      481 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
      541 taacaaaata ttaacgctta caatttcctg atgcggtatt ttctccttac gcatctgtgc
      601 ggtatttcac accgcatagg gtaataactg atataattaa attgaagctc taatttgtga
      661 gtttagtata catgcattta cttataatac agttttttag ttttgctggc cgcatcttct
      721 caaatatgct tcccagcctg cttttctgta acgttcaccc tctaccttag catcccttcc
      781 ctttgcaaat agtcctcttc caacaataat aatgtcagat cctgtagaga ccacatcatc
      841 cacggttcta tactgttgac ccaatgcgtc tcccttgtca tctaaaccca caccgggtgt
      901 cataatcaac caatcgtaac cttcatctct tccacccatg tctctttgag caataaagcc
      961 gataacaaaa tctttgtcgc tcttcgcaat gtcaacagta cccttagtat attctccagt
     1021 agatagggag cccttgcatg acaattctgc taacatcaaa aggcctctag gttcctttgt
     1081 tacttcttct gccgcctgct tcaaaccgct aacaatacct gggcccacca caccgtgtgc
     1141 attcgtaatg tctgcccatt ctgctattct gtatacaccc gcagagtact gcaatttgac
     1201 tgtattacca atgtcagcaa attttctgtc ttcgaagagt aaaaaattgt acttggcgga
     1261 taatgccttt agcggcttaa ctgtgccctc catggaaaaa tcagtcaaga tatccacatg
     1321 tgtttttagt aaacaaattt tgggacctaa tgcttcaact aactccagta attccttggt
     1381 ggtacgaaca tccaatgaag cacacaagtt tgtttgcttt tcgtgcatga tattaaatag
     1441 cttggcagca acaggactag gatgagtagc agcacgttcc ttatatgtag ctttcgacat
     1501 gatttatctt cgtttcctgc aggtttttgt tctgtgcagt tgggttaaga atactgggca
     1561 atttcatgtt tcttcaacac tacatatgcg tatatatacc aatctaagtc tgtgctcctt
     1621 ccttcgttct tccttctgtt cggagattac cgaatcaaaa aaatttcaag gaaaccgaaa
     1681 tcaaaaaaaa gaataaaaaa aaaatgatga attgaaaagg tggtatggtg cactctcagt
     1741 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac
     1801 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     1861 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     1921 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacggat
     1981 cgcttgcctg taacttacac gcgcctcgta tcttttaatg atggaataat ttgggaattt
     2041 actctgtgtt tatttatttt tatgttttgt atttggattt tagaaagtaa ataaagaagg
     2101 tagaagagtt acggaatgaa gaaaaaaaaa taaacaaagg tttaaaaaat ttcaacaaaa
     2161 agcgtacttt acatatatat ttattagaca agaaaagcag attaaataga tatacattcg
     2221 attaacgata agtaaaatgt aaaatcacag gattttcgtg tgtggtcttc tacacagaca
     2281 agatgaaaca attcggcatt aatacctgag agcaggaaga gcaagataaa aggtagtatt
     2341 tgttggcgat ccccctagag tcttttacat cttcggaaaa caaaaactat tttttcttta
     2401 atttcttttt ttactttcta tttttaattt atatatttat attaaaaaat ttaaattata
     2461 attattttta tagcacgtga tgaaaaggac ccaggtggca cttttcgggg aaatgtgcgc
     2521 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     2581 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     2641 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     2701 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     2761 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     2821 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     2881 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     2941 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     3001 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     3061 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     3121 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     3181 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     3241 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     3301 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     3361 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     3421 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     3481 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     3541 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     3601 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     3661 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     3721 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     3781 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     3841 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     3901 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     3961 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     4021 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     4081 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     4141 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     4201 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     4261 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     4321 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     4381 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     4441 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     4501 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     4561 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     4621 tttcacacag gaaacagcta tgaccatgat tacgccaagc tcgaaattaa ccctcactaa
     4681 agggaacaaa agctggtacc gggccccccc tcgaggtgac catggttcgt aacgaggcca
     4741 attcggcact cggatcggta acaataaagg gtcttacggg gccctgcttg atatccagtt
     4801 cgggcacggg atcatcctgc tctacacgtt tgtacgtttt ctcttccgac atgaccgtgc
     4861 ttgtagctgt tagctgtttc ttgataggct cttctgatac tacctcgctt taacaatgca
     4921 cctgtttgtc ttgttttgct gctcgaaaat aatcaagatc tgcttagaaa cgccaaactc
     4981 ttcttcgaca cgtgtcgaga aaaaaaacgc agcgaaaaaa aaaaaggtat atcatcacaa
     5041 tatacgatat aaatgggcag ggcaaccccg aatgatgtat attctcttct ggttccaagg
     5101 gtggcaaatg ttggtagaaa gcgtaattga agcaaagatc cgttaaataa ggccaactaa
     5161 cagaagcgct gaaagaggac aattggtgat taatctataa acgaggaact tcatgtcaag
     5221 agatgcacca attaaggctg acaaggatta tagccaaatt ttgaaggaag agtttcctaa
     5281 gatcgattcg ctcgctcaaa atgattgtaa ctctgcttta gaccaactgt tactgttgga
     5341 gaagaaaacc agacaagctt cagatctggc ctcctcgaaa gaagttttgg ccaagattgt
     5401 ggatctgcta gcatcaagga ataagtggga cgacctaaat gagcaattga ctctactctc
     5461 aaaaaagcat ggtcagttga aattgtcaat tcagtatatg atacaaaagg ttatggaata
     5521 tttgaaaagc tcgaaatctt tggatttaaa caccagaatt agtgtcattg aaactatcag
     5581 ggtggttaca gagaacaaaa tatttgtaga agtggaaaga gctagggtca ccaaagattt
     5641 ggtagaaatt aagaaagaag agggtaagat tgatgaagct gcagacatct tgtgtgagtt
     5701 acaggttgag acctatggct ccatggaaat gtctgagaaa attcagttta tattagagca
     5761 aatggaattg agtatattaa aaggtgatta ttcccaagcc acggtgcttt caagaaaaat
     5821 tctgaaaaaa acttttaaaa atccaaaata cgagtcattg aagctagaat attataatct
     5881 tctggtaaaa attagtttgc acaagagaga atacctagaa gttgcgcagt atctgcaaga
     5941 aatttatcaa acagacgcca ttaaatcaga tgaggctaag tggaaacctg ttttatcgca
     6001 cattgtatat ttcttagtcc tttcacctta cggcaattta caaaatgatt taattcacaa
     6061 aatccagaat gataacaacc tgaaaaaatt agaaagccaa gaatctttag taaaattgtt
     6121 tactacgaat gagttgatga gatggccaat tgttcaaaaa acctatgagc ccgtcttaaa
     6181 tgaggatgat ttggcatttg gtggagaagc taataagcat cactgggaag atttacaaaa
     6241 aagggtcatc gagcacaatt taagagtcat ttccgaatac tattccagaa ttactttact
     6301 aagattgaat gaattgctgg acctaacgga gagccagacg gaaacataca tcagtgattt
     6361 ggtaaaccag ggcatcatat acgctaaagt taatcgccca gccaaaatcg tgaattttga
     6421 aaaaccaaaa aactcaagcc aattattgaa cgaatggtca cataatgttg acgaactatt
     6481 agaacatata gaaacaatag gccatttaat tacaaaagag gaaatcatgc acggtctgca
     6541 agctaaatga agagattttc gagtaactgt cggattatct cagatccggt tttgatttta
     6601 aatctgtcac taatcgaaat gtgtatttac gtacgtgaga taaaattatc gagctccaat
     6661 tcgccctata gtgagtcgta ttacaattca ctggccgtcg ttttacaacg tcgtgactgg
     6721 gaaaaccctg gcgtta
//
