LOCUS       sequence-459454-        8576 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Z-tagged LRRK2 KW using Baculoviruses.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8576)
  AUTHORS   Raig ND, Surridge KJ, Sanz-Murillo M, Dederer V, Kramer A, Schwalm 
            MP, Elson L, Chatterjee D, Mathea S, Hanke T, Leschziner AE, 
            Reck-Peterson SL, Knapp S
  TITLE     Type-II kinase inhibitors that target Parkinson's Disease-associated
            LRRK2.
  JOURNAL   bioRxiv [Preprint]. 2025 Feb 13:2024.09.17.613365. doi: 
            10.1101/2024.09.17.613365.
  PUBMED    39554022
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: "bioRxiv 
            [Preprint]. 2025 Feb 13:2024.09.17.613365. doi: 
            10.1101/2024.09.17.613365."
COMMENT     Sequence Label: pFB-EPCP5
FEATURES             Location/Qualifiers
     source          1..8576
                     /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             4103..4120
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4130..4300
                     /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             5870..5890
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     polyA_signal    8066..8200
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8103..8122)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8157..8176
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  8229..8394
                     /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 gcggaattca aaggcctacg tcgacgagct
     4081 cactagtcgc ggccgcatgg gacaccacca ccaccaccac tccagcggag tggacaacaa
     4141 gttcaacaag gaacgccgta gggctagacg cgagatccgt cacctgccca acctgaacag
     4201 ggagcagcgt agggccttca tcagatccct gcgcgacgac ccatcacagt ccgctaacct
     4261 gctggctgaa gccaagaagc tgaacgacgc ccagcctaag ggaacttccg gtacctccgg
     4321 tggttccatg aagaaggctg tgccctacaa ccgtatgaag ctgatgatcg tcggtaacac
     4381 cggtagcggc aagaccactc tgctgcagca gctgatgaag accaagaagt ccgacctggg
     4441 catgcagagc gctactgtcg gaatcgacgt gaaggactgg cctatccaga tccgcgacaa
     4501 gagaaagcgc gacctggtcc tgaacgtgtg ggacttcgcc ggccgtgagg aattctactc
     4561 tacccacccc cacttcatga ctcagagggc tctgtacctg gccgtgtacg acctgtcaaa
     4621 gggacaggct gaggtcgacg ccatgaagcc ctggctgttc aacatcaagg ctagagcctc
     4681 ttcatcccca gtcatcctgg tgggtactca cctggacgtg tccgacgaaa agcagcgcaa
     4741 ggcttgcatg agcaagatca ccaaggagct gctgaacaag cgtggcttcc cagctatcag
     4801 ggactaccac ttcgtcaacg ccaccgagga atccgacgct ctggccaagc tgagaaagac
     4861 tatcatcaac gagagcctga acttcaagat ccgcgaccag ctggtggtcg gccagctgat
     4921 ccctgactgc tacgtggagc tggaaaagat catcctgtcc gagcgtaaga acgtccccat
     4981 cgagttccca gtgatcgacc gtaagaggct gctgcagctg gtcagggaaa accagctgca
     5041 gctggacgag aacgaactgc cacacgctgt ccacttcctg aacgagtctg gagtgctgct
     5101 gcacttccag gacccagccc tgcagctgtc agacctgtac ttcgtggaac ctaagtggct
     5161 gtgcaagatc atggcccaga tcctgaccgt caaggtggaa ggttgcccca agcacccaaa
     5221 gggcatcatc tctagacgcg acgtcgagaa gttcctgtca aagaagcgta agttccctaa
     5281 gaactacatg tcccagtact tcaagctgct ggagaagttc cagatcgctc tgccaatcgg
     5341 cgaggaatac ctgctggtgc ctagctctct gtccgaccac agaccagtca tcgaactgcc
     5401 tcactgcgaa aacagcgaga tcatcatccg cctgtacgag atgccttact tccccatggg
     5461 tttctggagc cgtctgatca acaggctgct ggaaatctcc ccctacatgc tcagcggcag
     5521 agagagggct ctgaggccaa acaggatgta ctggcgtcag ggaatctacc tgaactggtc
     5581 ccccgaagcc tactgcctgg tcggtagcga agtgctggac aaccaccccg agtccttcct
     5641 gaagatcact gtgccaagct gcaggaaggg atgcatcctg ctgggtcaag tggtcgacca
     5701 catcgactct ctgatggagg aatggttccc aggtctgctg gagatcgaca tctgcggaga
     5761 gggtgaaacc ttgctgaaga agtgggctct gtactcattc aacgacggcg aggaacacca
     5821 gaagatcctg ctggacgacc tgatgaagaa ggccgaggaa ggagacaccg agaacctgta
     5881 cttccagtcc cgtctgacta tccctatctc tcagatcgct cctgacctga tcctggctga
     5941 cctgccaagg aacatcatgc tgaacaacga cgagctggaa ttcgagcagg ctcctgaatt
     6001 cctgctgggc gacggatcat tcggttctgt gtacagggct gcttacgagg gtgaagaagt
     6061 cgccgtgaag atcttcaaca agcacaccag cctgagactg ctgcgccagg aactggtggt
     6121 cctgtgccac ctgcaccacc ctagcctgat ctctctgctg gctgccggca tccgtcccag
     6181 gatgctggtc atggagctgg cttcaaaggg atccctggac cgtctgctgc agcaggacaa
     6241 ggcctctctg accagaactc tgcaacacag gatcgctctg cacgtggctg acggactgag
     6301 atacctgcac tcagctatga tcatctaccg cgacctgaag cctcacaacg tcctgctgtt
     6361 caccctgtac cccaacgctg ccatcatcgc taagatcgcc gactacggaa tcgcccagta
     6421 ctgctgccgt atgggtatca agacctccga aggaactccc ggtttcagag ctccagaggt
     6481 cgcccgcggt aacgtgatct acaaccagca ggctgacgtg tacagcttcg gcctgctgct
     6541 gtacgacatc ctgaccactg gtggcaggat cgtcgaaggt ctgaagttcc ctaacgagtt
     6601 cgacgaactg gagatccagg gcaagctgcc tgaccccgtg aaggaatacg gatgcgctcc
     6661 atggcctatg gtcgagaagc tgatcaagca gtgcctgaag gaaaaccctc aggagcgccc
     6721 cacctctgct caggtgttcg acatcctgaa ctcagccgaa ctggtctgcc tgactcgtag
     6781 gatcctgctg cctaagaacg tcatcgtgga gtgcatggtg gctacccacc acaacagcag
     6841 aaacgcctct atctggctgg gttgcggcca caccgaccgt ggtcagctgt ccttcctgga
     6901 cctgaacacc gagggataca cttctgagga agtcgctgac tcacgcatcc tctgcctggc
     6961 tctggtccac ctgccagtgg aaaaggagtc atggatcgtg tccggcaccc agagcggaac
     7021 tctgctggtc atcaacaccg aagacggcaa gaagcgccac actctggaga agatgaccga
     7081 ctctgtgact tgcctgtact gcaacagctt ctctaagcag tcaaagcaga agaacttcct
     7141 gctggtgggt accgctgacg gcaagctggc catcttcgag gacaagactg tcaagctgaa
     7201 gggtgctgcc ccactgaaga tcctgaacat cggcaacgtg tccacccctc tgatgtgcct
     7261 gtcagaatcc accaacagca ctgagcgtaa cgtcatgtgg ggaggttgcg gtaccaagat
     7321 cttcagcttc tctaacgact tcactatcca gaagctgatc gagaccagga cttctcagct
     7381 gttctcatac gctgccttct cagactccaa catcatcacc gtggtcgtgg acactgctct
     7441 gtacatcgcc aagcagaact cccccgtcgt ggaagtgtgg gacaagaaga ctgagaagct
     7501 gtgcggcctg atcgactgcg tccacttcct gagagaggtc atggtgaagg aaaacaagga
     7561 gagcaagcac aagatgagct actctggacg cgtgaagacc ctgtgcctgc agaagaacac
     7621 cgccctgtgg atcggtactg gcggaggtca catcctgctg ctggacctgt ctactagacg
     7681 cctgatccgt gtcatctaca acttctgcaa ctccgtgcgc gtgatgatga ccgctcagct
     7741 gggctcactg aagaacgtca tgctggtgct gggatacaac agaaagaaca ccgaaggtac
     7801 tcagaagcag aaggagatcc agtcatgcct gactgtctgg gacatcaacc tgccacacga
     7861 agtgcagaac ctggaaaagc acatcgaggt ccgcaaggaa ctggctgaga agatgcgtag
     7921 gacctctgtg gagtgaaagc ttgtcgagaa gtactagagg atcataatca gccataccac
     7981 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
     8041 taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata
     8101 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     8161 tttgtccaaa ctcatcaatg tatcttatca tgtctggatc tgatcactgc ttgagcctag
     8221 gagatccgaa ccagataagt gaaatctagt tccaaactat tttgtcattt ttaattttcg
     8281 tattagctta cgacgctaca cccagttccc atctattttg tcactcttcc ctaaataatc
     8341 cttaaaaact ccatttccac ccctcccagt tcccaactat tttgtccgcc cacagcgggg
     8401 catttttctt cctgttatgt ttttaatcaa acatcctgcc aactccatgt gacaaaccgt
     8461 catcttcggc tactttttct ctgtcacaga atgaaaattt ttctgtcatc tcttcgttat
     8521 taatgtttgt aattgactga atatcaacgc ttatttgcag cctgaatggc gaatgg
//
