LOCUS       sequence-426726-        7071 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of His6-E3(1)-CUbo-TwinStrep in insect cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7071)
  AUTHORS   Renz C, Asimaki E, Meister C, Albanese V, Petriukov K, Krapoth NC, 
            Wegmann S, Wollscheid HP, Wong RP, Fulzele A, Chen JX, Leon S, 
            Ulrich HD
  TITLE     Ubiquiton-An inducible, linkage-specific polyubiquitylation tool.
  JOURNAL   Mol Cell. 2024 Jan 18;84(2):386-400.e11. doi: 
            10.1016/j.molcel.2023.11.016. Epub 2023 Dec 15.
  PUBMED    38103558
REFERENCE   2  (bases 1 to 7071)
  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: 
            "10.1016/j.molcel.2023.11.016"; journalName: "Mol Cell"; date: 
            "2024-01-18- 18"; volume: "84"; issue: "2"; pages: "386-400.e11"
COMMENT     Sequence Label: pFastBac1-His6-E3(1)-CUbo-TwinStrep
FEATURES             Location/Qualifiers
     source          1..7071
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      2..456
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(89..108)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     298..319
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        482..586
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(587..655,656..1447)
                     /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(805..824)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1618..2206
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2107..2126
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2360..2377
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  2511..2735
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(2802..3335)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3524..3529,3530..3546,3547..3552))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     promoter        3904..3995
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     3920..3937
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             4065..4082
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5859..6179
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             join(6327..6332,6333..6356,6357..6392,6393..6416)
                     /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    6561..6695
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6598..6617)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6652..6671
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  6724..6889
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
       61 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
      121 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
      181 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
      241 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt cttaatagtg
      301 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
      361 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      421 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
      481 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
      541 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
      601 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
      661 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
      721 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
      781 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
      841 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
      901 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
      961 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     1021 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     1081 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     1141 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     1201 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     1261 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     1321 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     1381 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     1441 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     1501 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     1561 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     1621 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     1681 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     1741 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     1801 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     1861 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     1921 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     1981 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     2041 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     2101 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     2161 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     2221 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     2281 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     2341 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg
     2401 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atagaccagc cgcgtaacct
     2461 ggcaaaatcg gttacggttg agtaataaat ggatgccctg cgtaagcggg tgtgggcgga
     2521 caataaagtc ttaaactgaa caaaatagat ctaaactatg acaataaagt cttaaactag
     2581 acagaatagt tgtaaactga aatcagtcca gttatgctgt gaaaaagcat actggacttt
     2641 tgttatggct aaagcaaact cttcattttc tgaagtgcaa attgcccgtc gtattaaaga
     2701 ggggcgtggc caagggcatg gtaaagacta tattcgcggc gttgtgacaa tttaccgaac
     2761 aactccgcgg ccgggaagcc gatctcggct tgaacgaatt gttaggtggc ggtacttggg
     2821 tcgatatcaa agtgcatcac ttcttcccgt atgcccaact ttgtatagag agccactgcg
     2881 ggatcgtcac cgtaatctgc ttgcacgtag atcacataag caccaagcgc gttggcctca
     2941 tgcttgagga gattgatgag cgcggtggca atgccctgcc tccggtgctc gccggagact
     3001 gcgagatcat agatatagat ctcactacgc ggctgctcaa acttgggcag aacgtaagcc
     3061 gcgagagcgc caacaaccgc ttcttggtcg aaggcagcaa gcgcgatgaa tgtcttacta
     3121 cggagcaagt tcccgaggta atcggagtcc ggctgatgtt gggagtaggt ggctacgtct
     3181 ccgaactcac gaccgaaaag atcaagagca gcccgcatgg atttgacttg gtcagggccg
     3241 agcctacatg tgcgaatgat gcccatactt gagccaccta actttgtttt agggcgactg
     3301 ccctgctgcg taacatcgtt gctgctgcgt aacatcgttg ctgctccata acatcaaaca
     3361 tcgacccacg gcgtaacgcg cttgctgctt ggatgcccga ggcatagact gtacaaaaaa
     3421 acagtcataa caagccatga aaaccgccac tgcgccgtta ccaccgctgc gttcggtcaa
     3481 ggttctggac cagttgcgtg agcgcatacg ctacttgcat tacagtttac gaaccgaaca
     3541 ggcttatgtc aactgggttc gtgccttcat ccgtttccac ggtgtgcgtc acccggcaac
     3601 cttgggcagc agcgaagtcg aggcatttct gtcctggctg gcgaacgagc gcaaggtttc
     3661 ggtctccacg catcgtcagg cattggcggc cttgctgttc ttctacggca aggtgctgtg
     3721 cacggatctg ccctggcttc aggagatcgg aagacctcgg ccgtcgcggc gcttgccggt
     3781 ggtgctgacc ccggatgaag tggttcgcat cctcggtttt ctggaaggcg agcatcgttt
     3841 gttcgcccag gactctagct atagttctag tggttggcta cgtatactcc ggaatattaa
     3901 tagatcatgg agataattaa aatgataacc atctcgcaaa taaataagta ttttactgtt
     3961 ttcgtaacag ttttgtaata aaaaaaccta taaatattcc ggattattca taccgtccca
     4021 ccatcgggcg cggatcccgg tccgaagcgc gcatgggcag cagccatcat catcatcatc
     4081 acagcagcgg ccagattttc gtcaagactt tgaccggtaa aaccataaca ttggaagttg
     4141 aatcttccga taccatcgac aacgttaagt cgcgtattca agacaaggaa ggcattccac
     4201 ctgatcaaca aagattgatc tttgccggta gacagctcga ggacggtaga acgctgtctg
     4261 attacaacat tcagcgtgag tccaccttac atcttgtgct aaggctacgt ggcctcatgc
     4321 agattttcgt caagactttg accggtaaaa ccataacatt ggaagttgaa tcttccgata
     4381 ccatcgacaa cgttaagtcg cgtattcaag acaaggaagg cattccacct gatcaacaaa
     4441 gattgatctt tgccggtaga cagctcgagg acggtagaac gctgtctgat tacaacattc
     4501 agcgtgagtc caccttacat cttgtgctaa ggctacgtgg cctcgcttct ggtgctggcg
     4561 gttctgaagg tggcggatct gaaggtggca catctggcgc tacaagatct gcttctggtg
     4621 ctggtggatc tgaaggtgga ggttctgaag gtggtacctc tggtgctacc tgtgctgttt
     4681 gtggttgggc tttgccacac aacagaatgc aagctttgac ctcttgtgaa tgtaccatct
     4741 gtccagactg tttcagacaa cacttcacca tcgctttgaa ggaaaagcac atcaccgaca
     4801 tggtttgtcc agcttgtggt agaccagact tgaccgacga cacccaattg ttgtcttact
     4861 tctctacctt ggacatccaa ttgagagaat ctttggaacc agacgcttac gctttgttcc
     4921 acaagaagtt gaccgaaggt gttttgatga gagacccaaa gttcttgtgg tgtgctcaat
     4981 gttctttcgg tttcatctac gaaagagaac aattggaagc tacctgtcca caatgtcacc
     5041 aaaccttctg tgttagatgt aagagacaat gggaagaaca acacagaggt agatcatgtg
     5101 aagacttcca aaactggaag agaatgaacg acccagaata ccaagctcaa ggtttggcta
     5161 tgtacttgca agaaaacggt atcgactgtc caaagtgtaa gttctcttac gctttggcta
     5221 gaggtggttg tatgcacttc cactgtaccc aatgtagaca ccaattctgt tctggttgtt
     5281 acaacgcttt ctacgctaag aacaagtgtc cagaaccaaa ctgtagagtt aagaagtctt
     5341 tgcacggtca ccacccaaga gactgtttgt tctacttgag agactggacc gctttgagat
     5401 tgcaaaagtt gttgcaagac aacaacgtta tgttcaacac cgaaccacca gctggtgcta
     5461 gagctgttcc aggtggtggt tgtagagtta tcgaacaaaa ggaagttcca aacggtttga
     5521 gagacgaagc ttgtggtaag gaaaccccag ctggttacgc tggtttgtgt caagctcact
     5581 acaaggaata cttggtttct ttgatcaacg ctcactcttt ggacccagct accttgtacg
     5641 aagttgaaga attggaaacc gctaccgaaa gatacttgca cgttagacca caaccattgg
     5701 ctggtgaaga cccaccagct taccaagcta gattgttgca aaagttgacc gaagaagttc
     5761 cattgggtca atctatccca agaagaagaa agggatccgc ttctggtgct ggtggatctg
     5821 aaggtggagg ttctgaaggt ggtacctctg gtgctagcgg tgttcaagtt gaaaccatct
     5881 ctccaggtga cggtagaacc ttcccaaaga gaggtcaaac ctgtgttgtt cactacaccg
     5941 gtatgttgga agacggtaag aagttcgact cttctagaga cagaaacaag ccattcaagt
     6001 tcatgttggg taagcaagaa gttatcagag gttgggaaga aggtgttgct caaatgtctg
     6061 ttggtcaaag agctaagttg accatctctc cagactacgc ttacggtgct accggtcacc
     6121 caggtattat cccaccacac gctaccttgg ttttcgacgt tgaattgttg aagttggaag
     6181 ggtcgaccgg tatcccacca gaccaacaaa gattgatctt cgctggtaag caattggaag
     6241 acggtagaac cttgtctgac tacaacatcc aaaaggaatc taccttgcac ttggttttga
     6301 gattgagagg tgttttaatt aatggtagtg catggagcca tccgcagttt gaaaaaggtg
     6361 gtggtagcgg tggtggttca ggtggtagtg catggtcaca ccctcagttt gagaaatagg
     6421 catgcggtac caagcttgtc gagaagtact agaggatcat aatcagccat accacatttg
     6481 tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa
     6541 tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca
     6601 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     6661 ccaaactcat caatgtatct tatcatgtct ggatctgatc actgcttgag cctaggagat
     6721 ccgaaccaga taagtgaaat ctagttccaa actattttgt catttttaat tttcgtatta
     6781 gcttacgacg ctacacccag ttcccatcta ttttgtcact cttccctaaa taatccttaa
     6841 aaactccatt tccacccctc ccagttccca actattttgt ccgcccacag cggggcattt
     6901 ttcttcctgt tatgttttta atcaaacatc ctgccaactc catgtgacaa accgtcatct
     6961 tcggctactt tttctctgtc acagaatgaa aatttttctg tcatctcttc gttattaatg
     7021 tttgtaattg actgaatatc aacgcttatt tgcagcctga atggcgaatg g
//
