LOCUS       sequence-450970-        8576 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Z-tagged LRRK2 RCKW using Baculoviruses.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8576)
  AUTHORS   Dederer V, Sanz Murillo M, Karasmanis EP, Hatch KS, Chatterjee D, 
            Preuss F, Abdul Azeez KR, Nguyen LV, Galicia C, Dreier B, Pluckthun 
            A, Versees W, Mathea S, Leschziner AE, Reck-Peterson SL, Knapp S
  TITLE     A designed ankyrin-repeat protein that targets Parkinson's 
            disease-associated LRRK2.
  JOURNAL   J Biol Chem. 2024 Jul;300(7):107469. doi: 10.1016/j.jbc.2024.107469.
            Epub 2024 Jun 12.
  PUBMED    38876305
REFERENCE   2  (bases 1 to 8576)
  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: "J Biol 
            Chem."; date: "2024-07"; volume: "300(7):107469. doi"; pages: " 
            10.1016/j.jbc.2024.107469. Epub 2024 Jun 12"
COMMENT     Sequence Label: pFB-EPCP1
FEATURES             Location/Qualifiers
     source          1..8576
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(1532..1552)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             complement(1559..1729)
                     /codon_start=1
                     /product="Z domain of staphylococcal protein A, engineered 
                     to contain multiple positive charges (Hedhammar and Hober, 
                     2007)"
                     /label=Z-basic tag
                     /note="useful for purifying tagged proteins by 
                     cation-exchange chromatography"
                     /translation="VDNKFNKERRRARREIRHLPNLNREQRRAFIRSLRDDPSQSANLL
                     AEAKKLNDAQPK"
     CDS             complement(1739..1756)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        complement(1864..1955)
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     complement(1922..1939)
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     promoter        join(2307..2312,2313..2329,2330..2335)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             2524..3057
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     mobile_element  3124..3348
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     primer_bind     complement(3482..3499)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3653..4241)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3733..3752)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4412..5203,5204..5272))
                     /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     5035..5054
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5273..5377)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5403..5857)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5540..5561)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5751..5770
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     mobile_element  6041..6206
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     polyA_signal    6235..6369
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6259..6278)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6313..6332
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
ORIGIN
        1 cctcggcctt cttcatcagg tcgtccagca ggatcttctg gtgttcctcg ccgtcgttga
       61 atgagtacag agcccacttc ttcagcaagg tttcaccctc tccgcagatg tcgatctcca
      121 gcagacctgg gaaccattcc tccatcagag agtcgatgtg gtcgaccact tgacccagca
      181 ggatgcatcc cttcctgcag cttggcacag tgatcttcag gaaggactcg gggtggttgt
      241 ccagcacttc gctaccgacc aggcagtagg cttcggggga ccagttcagg tagattccct
      301 gacgccagta catcctgttt ggcctcagag ccctctctct gccgctgagc atgtaggggg
      361 agatttccag cagcctgttg atcagacggc tccagaaacc catggggaag taaggcatct
      421 cgtacaggcg gatgatgatc tcgctgtttt cgcagtgagg cagttcgatg actggtctgt
      481 ggtcggacag agagctaggc accagcaggt attcctcgcc gattggcaga gcgatctgga
      541 acttctccag cagcttgaag tactgggaca tgtagttctt agggaactta cgcttctttg
      601 acaggaactt ctcgacgtcg cgtctagaga tgatgccctt tgggtgcttg gggcaacctt
      661 ccaccttgac ggtcaggatc tgggccatga tcttgcacag ccacttaggt tccacgaagt
      721 acaggtctga cagctgcagg gctgggtcct ggaagtgcag cagcactcca gactcgttca
      781 ggaagtggac agcgtgtggc agttcgttct cgtccagctg cagctggttt tccctgacca
      841 gctgcagcag cctcttacgg tcgatcactg ggaactcgat ggggacgttc ttacgctcgg
      901 acaggatgat cttttccagc tccacgtagc agtcagggat cagctggccg accaccagct
      961 ggtcgcggat cttgaagttc aggctctcgt tgatgatagt ctttctcagc ttggccagag
     1021 cgtcggattc ctcggtggcg ttgacgaagt ggtagtccct gatagctggg aagccacgct
     1081 tgttcagcag ctccttggtg atcttgctca tgcaagcctt gcgctgcttt tcgtcggaca
     1141 cgtccaggtg agtacccacc aggatgactg gggatgaaga ggctctagcc ttgatgttga
     1201 acagccaggg cttcatggcg tcgacctcag cctgtccctt tgacaggtcg tacacggcca
     1261 ggtacagagc cctctgagtc atgaagtggg ggtgggtaga gtagaattcc tcacggccgg
     1321 cgaagtccca cacgttcagg accaggtcgc gctttctctt gtcgcggatc tggataggcc
     1381 agtccttcac gtcgattccg acagtagcgc tctgcatgcc caggtcggac ttcttggtct
     1441 tcatcagctg ctgcagcaga gtggtcttgc cgctaccggt gttaccgacg atcatcagct
     1501 tcatacggtt gtagggcaca gccttcttca tggactggaa gtacaggttc tcagttccct
     1561 taggctgggc gtcgttcagc ttcttggctt cagccagcag gttagcggac tgtgatgggt
     1621 cgtcgcgcag ggatctgatg aaggccctac gctgctccct gttcaggttg ggcaggtgac
     1681 ggatctcgcg tctagcccta cggcgttcct tgttgaactt gttgtccact ccgctggagt
     1741 ggtggtggtg gtggtgtccc atgcggccgc gactagtgag ctcgtcgacg taggcctttg
     1801 aattccgcgc gcttcggacc gggatccgcg cccgatggtg ggacggtatg aataatccgg
     1861 aatatttata ggttttttta ttacaaaact gttacgaaaa cagtaaaata cttatttatt
     1921 tgcgagatgg ttatcatttt aattatctcc atgatctatt aatattccgg agtatacgta
     1981 gccaaccact agaactatag ctagagtcct gggcgaacaa acgatgctcg ccttccagaa
     2041 aaccgaggat gcgaaccact tcatccgggg tcagcaccac cggcaagcgc cgcgacggcc
     2101 gaggtcttcc gatctcctga agccagggca gatccgtgca cagcaccttg ccgtagaaga
     2161 acagcaaggc cgccaatgcc tgacgatgcg tggagaccga aaccttgcgc tcgttcgcca
     2221 gccaggacag aaatgcctcg acttcgctgc tgcccaaggt tgccgggtga cgcacaccgt
     2281 ggaaacggat gaaggcacga acccagttga cataagcctg ttcggttcgt aaactgtaat
     2341 gcaagtagcg tatgcgctca cgcaactggt ccagaacctt gaccgaacgc agcggtggta
     2401 acggcgcagt ggcggttttc atggcttgtt atgactgttt ttttgtacag tctatgcctc
     2461 gggcatccaa gcagcaagcg cgttacgccg tgggtcgatg tttgatgtta tggagcagca
     2521 acgatgttac gcagcagcaa cgatgttacg cagcagggca gtcgccctaa aacaaagtta
     2581 ggtggctcaa gtatgggcat cattcgcaca tgtaggctcg gccctgacca agtcaaatcc
     2641 atgcgggctg ctcttgatct tttcggtcgt gagttcggag acgtagccac ctactcccaa
     2701 catcagccgg actccgatta cctcgggaac ttgctccgta gtaagacatt catcgcgctt
     2761 gctgccttcg accaagaagc ggttgttggc gctctcgcgg cttacgttct gcccaagttt
     2821 gagcagccgc gtagtgagat ctatatctat gatctcgcag tctccggcga gcaccggagg
     2881 cagggcattg ccaccgcgct catcaatctc ctcaagcatg aggccaacgc gcttggtgct
     2941 tatgtgatct acgtgcaagc agattacggt gacgatcccg cagtggctct ctatacaaag
     3001 ttgggcatac gggaagaagt gatgcacttt gatatcgacc caagtaccgc cacctaacaa
     3061 ttcgttcaag ccgagatcgg cttcccggcc gcggagttgt tcggtaaatt gtcacaacgc
     3121 cgcgaatata gtctttacca tgcccttggc cacgcccctc tttaatacga cgggcaattt
     3181 gcacttcaga aaatgaagag tttgctttag ccataacaaa agtccagtat gctttttcac
     3241 agcataactg gactgatttc agtttacaac tattctgtct agtttaagac tttattgtca
     3301 tagtttagat ctattttgtt cagtttaaga ctttattgtc cgcccacacc cgcttacgca
     3361 gggcatccat ttattactca accgtaaccg attttgccag gttacgcggc tggtctatgc
     3421 ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc tcttccgctt
     3481 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     3541 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     3601 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     3661 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     3721 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     3781 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     3841 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     3901 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     3961 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     4021 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     4081 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     4141 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     4201 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     4261 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     4321 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     4381 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     4441 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     4501 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     4561 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     4621 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     4681 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     4741 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     4801 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     4861 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     4921 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     4981 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     5041 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     5101 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     5161 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     5221 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     5281 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     5341 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     5401 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     5461 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     5521 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaagaa cgtggactcc
     5581 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc
     5641 taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc
     5701 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa
     5761 gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc
     5821 acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt caggctgcaa
     5881 ataagcgttg atattcagtc aattacaaac attaataacg aagagatgac agaaaaattt
     5941 tcattctgtg acagagaaaa agtagccgaa gatgacggtt tgtcacatgg agttggcagg
     6001 atgtttgatt aaaaacataa caggaagaaa aatgccccgc tgtgggcgga caaaatagtt
     6061 gggaactggg aggggtggaa atggagtttt taaggattat ttagggaaga gtgacaaaat
     6121 agatgggaac tgggtgtagc gtcgtaagct aatacgaaaa ttaaaaatga caaaatagtt
     6181 tggaactaga tttcacttat ctggttcgga tctcctaggc tcaagcagtg atcagatcca
     6241 gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa
     6301 tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat
     6361 aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg
     6421 gaggtttttt aaagcaagta aaacctctac aaatgtggta tggctgatta tgatcctcta
     6481 gtacttctcg acaagctttc actccacaga ggtcctacgc atcttctcag ccagttcctt
     6541 gcggacctcg atgtgctttt ccaggttctg cacttcgtgt ggcaggttga tgtcccagac
     6601 agtcaggcat gactggatct ccttctgctt ctgagtacct tcggtgttct ttctgttgta
     6661 tcccagcacc agcatgacgt tcttcagtga gcccagctga gcggtcatca tcacgcgcac
     6721 ggagttgcag aagttgtaga tgacacggat caggcgtcta gtagacaggt ccagcagcag
     6781 gatgtgacct ccgccagtac cgatccacag ggcggtgttc ttctgcaggc acagggtctt
     6841 cacgcgtcca gagtagctca tcttgtgctt gctctccttg ttttccttca ccatgacctc
     6901 tctcaggaag tggacgcagt cgatcaggcc gcacagcttc tcagtcttct tgtcccacac
     6961 ttccacgacg ggggagttct gcttggcgat gtacagagca gtgtccacga ccacggtgat
     7021 gatgttggag tctgagaagg cagcgtatga gaacagctga gaagtcctgg tctcgatcag
     7081 cttctggata gtgaagtcgt tagagaagct gaagatcttg gtaccgcaac ctccccacat
     7141 gacgttacgc tcagtgctgt tggtggattc tgacaggcac atcagagggg tggacacgtt
     7201 gccgatgttc aggatcttca gtggggcagc acccttcagc ttgacagtct tgtcctcgaa
     7261 gatggccagc ttgccgtcag cggtacccac cagcaggaag ttcttctgct ttgactgctt
     7321 agagaagctg ttgcagtaca ggcaagtcac agagtcggtc atcttctcca gagtgtggcg
     7381 cttcttgccg tcttcggtgt tgatgaccag cagagttccg ctctgggtgc cggacacgat
     7441 ccatgactcc ttttccactg gcaggtggac cagagccagg cagaggatgc gtgagtcagc
     7501 gacttcctca gaagtgtatc cctcggtgtt caggtccagg aaggacagct gaccacggtc
     7561 ggtgtggccg caacccagcc agatagaggc gtttctgctg ttgtggtggg tagccaccat
     7621 gcactccacg atgacgttct taggcagcag gatcctacga gtcaggcaga ccagttcggc
     7681 tgagttcagg atgtcgaaca cctgagcaga ggtggggcgc tcctgagggt tttccttcag
     7741 gcactgcttg atcagcttct cgaccatagg ccatggagcg catccgtatt ccttcacggg
     7801 gtcaggcagc ttgccctgga tctccagttc gtcgaactcg ttagggaact tcagaccttc
     7861 gacgatcctg ccaccagtgg tcaggatgtc gtacagcagc aggccgaagc tgtacacgtc
     7921 agcctgctgg ttgtagatca cgttaccgcg ggcgacctct ggagctctga aaccgggagt
     7981 tccttcggag gtcttgatac ccatacggca gcagtactgg gcgattccgt agtcggcgat
     8041 cttagcgatg atggcagcgt tggggtacag ggtgaacagc aggacgttgt gaggcttcag
     8101 gtcgcggtag atgatcatag ctgagtgcag gtatctcagt ccgtcagcca cgtgcagagc
     8161 gatcctgtgt tgcagagttc tggtcagaga ggccttgtcc tgctgcagca gacggtccag
     8221 ggatcccttt gaagccagct ccatgaccag catcctggga cggatgccgg cagccagcag
     8281 agagatcagg ctagggtggt gcaggtggca caggaccacc agttcctggc gcagcagtct
     8341 caggctggtg tgcttgttga agatcttcac ggcgacttct tcaccctcgt aagcagccct
     8401 gtacacagaa ccgaatgatc cgtcgcccag caggaattca ggagcctgct cgaattccag
     8461 ctcgtcgttg ttcagcatga tgttccttgg caggtcagcc aggatcaggt caggagcgat
     8521 ctgagagata gggatagtca gacgaggctg gtctgggttc accagcaggt ctcctt
//
