LOCUS       sequence-432185-        7613 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Insect cell expression of human 20S CP proteasome subunit beta 
            type-1 (PSMB1), beta type-2 (PSMB2), beta type-3 (PSMB3), and beta 
            type-4 (PSMB4) with C-terminal Twin-Strep-tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7613)
  AUTHORS   Adolf F, Du J, Goodall EA, Walsh RM Jr, Rawson S, von Gronau S, 
            Harper JW, Hanna J, Schulman BA
  TITLE     Visualizing chaperone-mediated multistep assembly of the human 20S 
            proteasome.
  JOURNAL   Nat Struct Mol Biol. 2024 Apr 10. doi: 10.1038/s41594-024-01268-9.
  PUBMED    38600324
REFERENCE   2  (bases 1 to 7613)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat Struct 
            Mol Biol. 2024 Apr 10. doi: 10.1038/s41594-024-01268-9."
COMMENT     Sequence Label: pACEBac1-PSMB1-PSMB2-PSMB3-PSMB4-TEV-2xSTII
FEATURES             Location/Qualifiers
     source          1..7613
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    807..941
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1097..1188
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     polyA_signal    2039..2173
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        2329..2420
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     CDS             3277..3297
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             join(3304..3309,3310..3333,3334..3369,3370..3393)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="SAWSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     polyA_signal    3565..3699
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     mobile_element  3851..4016
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      4206..4794
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4695..4714
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     mobile_element  4929..5152
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(5219..5752)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(5941..5946,5947..5963,5964..5969))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     protein_bind    complement(6040..6073)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        6150..6241
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     polyA_signal    7200..7334
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        7490..7581
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
ORIGIN
        1 gcgcggatcc cggtccgaag cgcgcggaat tcatggagta cctcatcggt atccaaggcc
       61 ccgactatgt tcttgtcgcc tccgaccggg tggccgccag caatattgtc cagatgaagg
      121 acgatcatga caagatgttt aagatgagtg aaaagatatt actcctgtgt gttggagagg
      181 ctggagacac tgtacagttt gcagaatata ttcagaaaaa cgtgcaactt tataagatgc
      241 gaaatggata tgaattgtct cccacggcag cagctaactt cacacgccga aacctggctg
      301 actgtcttcg gagtcggacc ccatatcatg tgaacctcct cctggctggc tatgatgagc
      361 atgaagggcc agcgctgtat tacatggact acctggcagc cttggccaag gccccttttg
      421 cagcgcacgg ctatggtgcc ttcctgactc tcagtatcct cgaccgatac tacacaccga
      481 ctatctcacg tgagagggca gtggaactcc ttaggaaatg tctggaggag ctccagaaac
      541 gcttcatcct gaatctgcca accttcagtg ttcgaatcat tgacaaaaat ggcatccatg
      601 acctggataa catttccttc cccaaacagg gctcctaatg agcggccgct ttcgaatcta
      661 gagcctgcag tctcgacaag cttgtcgaga agtactagag gatcataatc agccatacca
      721 catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac
      781 ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat
      841 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
      901 gtttgtccaa actcatcaat gtatcttatc atgtctggat ctgatcactg cttgagccta
      961 gaagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag
     1021 caataactat cataaccatt taaataaacc taatgatgcc tgatgtttaa atcgacctac
     1081 tccggaatat taatagatca tggagataat taaaatgata accatctcgc aaataaataa
     1141 gtattttact gttttcgtaa cagttttgta ataaaaaaac ctataaatat tccggattat
     1201 tcataccgtc ccaccatcgg gcgcggatcc cggtccgaag cgcgcggaat tcatgtctat
     1261 tatgtcctat aacggagggg ccgtcatggc catgaagggg aagaactgtg tggccatcgc
     1321 tgcagacagg cgcttcggga tccaggccca gatggtgacc acggacttcc agaagatctt
     1381 tcccatgggt gaccggctgt acatcggtct ggccgggctc gccactgacg tccagacagt
     1441 tgcccagcgc ctcaagttcc ggctgaacct gtatgagttg aaggaaggtc ggcagatcaa
     1501 accttatacc ctcatgagca tggtggccaa cctcttgtat gagaaacggt ttggccctta
     1561 ctacactgag ccagtcattg ccgggttgga cccgaagacc tttaagccct tcatttgctc
     1621 tctagacctc atcggctgcc ccatggtgac tgatgacttt gtggtcagtg gcacctgcgc
     1681 cgaacaaatg tacggaatgt gtgagtccct ctgggagccc aacatggatc cggatcacct
     1741 gtttgaaacc atctcccaag ccatgctgaa tgctgtggac cgggatgcag tgtcaggcat
     1801 gggagtcatt gtccacatca tcgagaagga caaaatcacc accaggacac tgaaggcccg
     1861 aatggactaa tgagcggccg ctttcgaatc tagagcctgc agtctcgaca agcttgtcga
     1921 gaagtactag aggatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa
     1981 aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa
     2041 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     2101 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     2161 tcatgtctgg atctgatcac tgcttgagcc tagaagatcc ggctgctaac aaagcccgaa
     2221 aggaagctga gttggctgct gccaccgctg agcaataact atcataacca tttaaataaa
     2281 cggttcacat agcttagttt aaatcgacct actccggaat attaatagat catggagata
     2341 attaaaatga taaccatctc gcaaataaat aagtatttta ctgttttcgt aacagttttg
     2401 taataaaaaa acctataaat attccggatt attcataccg tcccaccatc gggcgcggat
     2461 cccggtccga agcgcgcgga attcatggaa gcgtttttgg ggtcgcggtc cggactttgg
     2521 gcggggggtc cggccccagg acagttttac cgcattccgt ccactcccga ttccttcatg
     2581 gatccggcgt ctgcacttta cagaggtcca atcacgcgga cccagaaccc catggtgacc
     2641 gggacctcag tcctcggcgt taagttcgag ggcggagtgg tgattgccgc agacatgctg
     2701 ggatcctacg gctccttggc tcgtttccgc aacatctctc gcattatgcg agtcaacaac
     2761 agtaccatgc tgggtgcctc tggcgactac gctgatttcc agtatttgaa gcaagttctc
     2821 ggccagatgg tgattgatga ggagcttctg ggagatggac acagctatag tcctagagct
     2881 attcattcat ggctgaccag ggccatgtac agccggcgct cgaagatgaa ccctttgtgg
     2941 aacaccatgg tcatcggagg ctatgctgat ggagagagct tcctcggtta tgtggacatg
     3001 cttggtgtag cctatgaagc cccttcgctg gccactggtt atggtgcata cttggctcag
     3061 cctctgctgc gagaagttct ggagaagcag ccagtgctaa gccagaccga ggcccgcgac
     3121 ttagtagaac gctgcatgcg agtgctgtac taccgagatg cccgttctta caaccggttt
     3181 caaaccgcca ctgtcaccga aaaaggtgtt gaaatagagg gaccattgtc tacagagacc
     3241 aactgggata ttgcccacat gatcagtggc tttgaagagg atctgtactt tcagtctgtc
     3301 gacagtgctt ggagccatcc acagtttgag aaaggtggag gctctggcgg gggtagcggg
     3361 ggcagtgcct ggtctcaccc acaattcgaa aagtgataag cggccgcttt cgaatctaga
     3421 gcctgcagtc tcgacaagct tgtcgagaag tactagagga tcataatcag ccataccaca
     3481 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     3541 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     3601 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     3661 ttgtccaaac tcatcaatgt atcttatcat gtctggatct gatcactgct tgagcctaga
     3721 agatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca
     3781 ataactatca taaccattta aataaacact gacattgact tggtttccta gggtataccc
     3841 atctaattgg aaccagataa gtgaaatcta gttccaaact attttgtcat ttttaatttt
     3901 cgtattagct tacgacgcta cacccagttc ccatctattt tgtcactctt ccctaaataa
     3961 tccttaaaaa ctccatttcc acccctccca gttcccaact attttgtccg cccacaaccg
     4021 gttgacttgg gtcaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     4081 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     4141 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     4201 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     4261 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     4321 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     4381 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     4441 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     4501 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     4561 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     4621 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     4681 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     4741 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     4801 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     4861 cctgcgttat cccctgattg acttgggtcg ctcttcctgt ggatgcgcag atgccctgcg
     4921 taagcgggtg tgggcggaca ataaagtctt aaactgaaca aaatagatct aaactatgac
     4981 aataaagtct taaactagac agaatagttg taaactgaaa tcagtccagt tatgctgtga
     5041 aaaagcatac tggacttttg ttatggctaa agcaaactct tcattttctg aagtgcaaat
     5101 tgcccgtcgt attaaagagg ggcgtggcca agggcatgta aagactatat tcgcggcgtt
     5161 gtgacaattt accgaacaac tccgcggccg ggaagccgat ctcggcttga acgaattgtt
     5221 aggtggcggt acttgggtcg atatcaaagt gcatcacttc ttcccgtatg cccaactttg
     5281 tatagagagc cactgcggga tcgtcaccgt aatctgcttg cacgtagatc acataagcac
     5341 caagcgcgtt ggcctcatgc ttgaggagat tgatgagcgc ggtggcaatg ccctgcctcc
     5401 ggtgctcgcc ggagactgcg agatcataga tatagatctc actacgcggc tgctcaaact
     5461 tgggcagaac gtaagccgcg agagcgccaa caaccgcttc ttggtcgaag gcagcaagcg
     5521 cgatgaatgt cttactacgg agcaagttcc cgaggtaatc ggagtccggc tgatgttggg
     5581 agtaggtggc tacgtctccg aactcacgac cgaaaagatc aagagcagcc cgcatggatt
     5641 tgacttggtc agggccgagc ctacatgtgc gaatgatgcc catacttgag ccacctaact
     5701 ttgttttagg gcgactgccc tgctgcgtaa catcgttgct gctgcgtaac atcgttgctg
     5761 ctccataaca tcaaacatcg acccacggcg taacgcgctt gctgcttgga tgcccgaggc
     5821 atagactgta caaaaaaaca gtcataacaa gccatgaaaa ccgccactgc gccgttacca
     5881 ccgctgcgtt cggtcaaggt tctggaccag ttgcgtgagc gcatacgcta cttgcattac
     5941 agtttacgaa ccgaacaggc ttatgtcaac tgggttcgtg ccttcatccg tttccacggt
     6001 gtgcgtcacc cggcaacctt gggcagcagc gaagtcgcca taacttcgta tagcatacat
     6061 tatacgaagt tatctgtaac tataacggtc ctaaggtagc gagtttaaac actagtatcg
     6121 attcgcgacc tactccggaa tattaataga tcatggagat aattaaaatg ataaccatct
     6181 cgcaaataaa taagtatttt actgttttcg taacagtttt gtaataaaaa aacctataaa
     6241 tattccggat tattcatacc gtcccaccat cgggcgcgga tcccggtccg aagcgcgcgg
     6301 aattcatgtt gtcctctaca gccatgtatt cggctcctgg cagagacttg gggatggaac
     6361 cgcacagagc cgcgggccct ttgcagctgc gattttcgcc ctacgttttc aacggaggta
     6421 ctatactggc aattgctgga gaagattttg caattgttgc ttctgatact cgattgagtg
     6481 aagggttttc aattcatacg cgggatagcc ccaaatgtta caaattaaca gacaaaacag
     6541 tcattggatg cagcggtttt catggagact gtcttacgct gacaaagatt attgaagcaa
     6601 gactaaagat gtataagcat tccaataata aggccatgac tacgggggca attgctgcaa
     6661 tgctgtctac aatcctgtat tcaaggcgct tctttccata ctatgtttac aacatcatcg
     6721 gtggacttga tgaagaagga aagggggctg tatacagctt tgatccagta gggtcttacc
     6781 agagagactc cttcaaggct ggaggctcag caagtgccat gctacagccc ctgcttgaca
     6841 accaggttgg ttttaagaac atgcagaatg tggagcatgt tccgctgtcc ttggacagag
     6901 cgatgcggct ggtgaaagat gtcttcattt ctgcggctga gagagatgtg tacactgggg
     6961 acgcactccg gatctgcata gtgaccaaag agggcatcag ggaggaaact gtttccttaa
     7021 ggaaggactg ataagcggcc gctttcgaat ctagagcctg cagtctcgac aagcttgtcg
     7081 agaagtacta gaggatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     7141 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     7201 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     7261 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     7321 atcatgtctg gatctgatca ctgcttgagc ctagaagatc cggctgctaa caaagcccga
     7381 aaggaagctg agttggctgc tgccaccgct gagcaataac tatcataacc atttaaataa
     7441 actggatact attgcacgtt taaatcgacc tactccggaa tattaataga tcatggagat
     7501 aattaaaatg ataaccatct cgcaaataaa taagtatttt actgttttcg taacagtttt
     7561 gtaataaaaa aacctataaa tattccggat tattcatacc gtcccaccat cgg
//
