LOCUS       sequence-452258-        4576 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of yeast Rpn10.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4576)
  AUTHORS   Beckwith R, Estrin E, Worden EJ, Martin A
  TITLE     Reconstitution of the 26S proteasome reveals functional asymmetries 
            in its AAA+ unfoldase.
  JOURNAL   Nat Struct Mol Biol. 2013 Oct;20(10):1164-72. doi: 
            10.1038/nsmb.2659. Epub 2013 Sep 8.
  PUBMED    24013205
REFERENCE   2  (bases 1 to 4576)
  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"; doi: "10.1038/nsmb.2659";
            journalName: "Nat Struct Mol Biol"; date: "2013-10"; volume: "20"; 
            issue: "10"; pages: "1164-72"
COMMENT     Sequence Label: pAM239 Rpn10
FEATURES             Location/Qualifiers
     source          1..4576
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(144..803)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     718..737
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(804..906)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(1298..1315)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1432..1977)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(1512..1531)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2202..3284)
                     /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     3246..3265
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3285..3362)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3408..3427
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3408..3426
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3427..3451
                     /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)."
     CDS             3501..3518
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4356..4400
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(4438..4456)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4452..4499
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 ataagcggct atttaacgac cctgccctga accgacgacc gggtcgaatt tgctttcgaa
       61 tttctgccat tcatccgctt attatcactt attcaggcgt agcaaccagg cgtttaaggg
      121 caccaataac tgccttaaaa aaattacgcc ccgccctgcc actcatcgca gtactgttgt
      181 aattcattaa gcattctgcc gacatggaag ccatcacaaa cggcatgatg aacctgaatc
      241 gccagcggca tcagcacctt gtcgccttgc gtataatatt tgcccatagt gaaaacgggg
      301 gcgaagaagt tgtccatatt ggccacgttt aaatcaaaac tggtgaaact cacccaggga
      361 ttggctgaga cgaaaaacat attctcaata aaccctttag ggaaataggc caggttttca
      421 ccgtaacacg ccacatcttg cgaatatatg tgtagaaact gccggaaatc gtcgtggtat
      481 tcactccaga gcgatgaaaa cgtttcagtt tgctcatgga aaacggtgta acaagggtga
      541 acactatccc atatcaccag ctcaccgtct ttcattgcca tacggaactc cggatgagca
      601 ttcatcaggc gggcaagaat gtgaataaag gccggataaa acttgtgctt atttttcttt
      661 acggtcttta aaaaggccgt aatatccagc tgaacggtct ggttataggt acattgagca
      721 actgactgaa atgcctcaaa atgttcttta cgatgccatt gggatatatc aacggtggta
      781 tatccagtga tttttttctc cattttagct tccttagctc ctgaaaatct cgataactca
      841 aaaaatacgc ccggtagtga tcttatttca ttatggtgaa agttggaacc tcttacgtgc
      901 cgatcaacgt ctcattttcg ccaaaagttg gcccagggct tcccggtatc aacagggaca
      961 ccaggattta tttattctgc gaagtgatct tccgtcacag gtatttattc ggcgcaaagt
     1021 gcgtcgggtg atgctgccaa cttactgatt tagtgtatga tggtgttttt gaggtgctcc
     1081 agtggcttct gtttctatca gctgtccctc ctgttcagct actgacgggg tggtgcgtaa
     1141 cggcaaaagc accgccggac atcagcgcta gcggagtgta tactggctta ctatgttggc
     1201 actgatgagg gtgtcagtga agtgcttcat gtggcaggag aaaaaaggct gcaccggtgc
     1261 gtcagcagaa tatgtgatac aggatatatt ccgcttcctc gctcactgac tcgctacgct
     1321 cggtcgttcg actgcggcga gcggaaatgg cttacgaacg gggcggagat ttcctggaag
     1381 atgccaggaa gatacttaac agggaagtga gagggccgcg gcaaagccgt ttttccatag
     1441 gctccgcccc cctgacaagc atcacgaaat ctgacgctca aatcagtggt ggcgaaaccc
     1501 gacaggacta taaagatacc aggcgtttcc ccctggcggc tccctcgtgc gctctcctgt
     1561 tcctgccttt cggtttaccg gtgtcattcc gctgttatgg ccgcgtttgt ctcattccac
     1621 gcctgacact cagttccggg taggcagttc gctccaagct ggactgtatg cacgaacccc
     1681 ccgttcagtc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggaaa
     1741 gacatgcaaa agcaccactg gcagcagcca ctggtaattg atttagagga gttagtcttg
     1801 aagtcatgcg ccggttaagg ctaaactgaa aggacaagtt ttggtgactg cgctcctcca
     1861 agccagttac ctcggttcaa agagttggta gctcagagaa ccttcgaaaa accgccctgc
     1921 aaggcggttt tttcgttttc agagcaagag attacgcgca gaccaaaacg atctcaagaa
     1981 gatcatctta ttaatcagat aaaatatttc tagatttcag tgcaatttat ctcttcaaat
     2041 gtagcacctg aagtcagccc catacgatat aagttgtaat tctcatgtta gtcatgcccc
     2101 gcgcccaccg gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg
     2161 gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt
     2221 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     2281 tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg
     2341 cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc
     2401 aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta
     2461 tcgtcgtatc ccactaccga gatgtccgca ccaacgcgca gcccggactc ggtaatggcg
     2521 cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc
     2581 tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt
     2641 tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga
     2701 cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg
     2761 accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg
     2821 ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca
     2881 gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg
     2941 agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc
     3001 accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc
     3061 gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt
     3121 tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc
     3181 cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag
     3241 acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat
     3301 tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg
     3361 tccgggatct cgacgctctc ccttatgcga ctcctgcatt aggaaattaa tacgactcac
     3421 tataggggaa ttgtgagcgg ataacaattc ccctgtagaa ataattttgt ttaactttaa
     3481 taaggagata taccatgggc catcaccatc accatcacgc cggcgtattg gaagctacag
     3541 tgttagtgat tgataattca gagtactctc gcaatggcga ctttcctagg acaaggtttg
     3601 aagcccagat cgactcggta gagttcatat ttcaagccaa gagaaacagc aatcctgaga
     3661 atacagtggg tcttatctct ggtgctggcg ccaaccccag ggtgttatct acgtttaccg
     3721 ccgagtttgg gaagattctt gctggactac acgacacgca gatcgagggt aagctgcata
     3781 tggccactgc gttgcagatc gctcagctga ctttgaagca tcgccagaat aaggtccaac
     3841 atcaaaggat tgtggcattt gtgtgtagcc caataagtga ttctcgagac gaattgatca
     3901 gattggcaaa aacactgaaa aagaataatg ttgccgtgga catcatcaat tttggagaga
     3961 ttgaacagaa cacggagctt ttggatgagt tcatagctgc agtgaacaac cctcaagaag
     4021 aaactagtca tttgcttact gtgacgcctg gccccagact gctgtacgag aacatcgcat
     4081 cttcacccat aattctcgaa gaaggatcct ccggtatggg cgcctttggt gggtctggcg
     4141 gtgattccga tgccaatggc acatttatgg acttcggggt agacccatca atggacccag
     4201 aactggcaat ggccttgcgt ctgtctatgg aagaagagca gcaaagacag gaaaggttaa
     4261 gacagcagca acaacaacaa gatcagcctg agcagtctga acagcctgaa caacaccaag
     4321 acaaatagaa gcttgcggcc gcactcgagt ctggtaaaga aaccgctgct gcgaaatttg
     4381 aacgccagca catggactcg tctactagcg cagcttaatt aacctaggct gctgccaccg
     4441 ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc
     4501 tgaaacctca ggcatttgag aagcacacgg tcacactgct tccggtagtc aataaaccgg
     4561 taaaccagca atagac
//
