LOCUS       sequence-366090-        6419 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  2xHis-LRRK2 Armadillo 1-552.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6419)
  AUTHORS   Vides EG, Adhikari A, Chiang CY, Lis P, Purlyte E, Limouse C, 
            Shumate JL, Spinola-Lasso E, Dhekne HS, Alessi DR, Pfeffer SR
  TITLE     A feed-forward pathway drives LRRK2 kinase membrane recruitment and 
            activation.
  JOURNAL   Elife. 2022 Sep 23;11:e79771. doi: 10.7554/eLife.79771.
  PUBMED    36149401
REFERENCE   2  (bases 1 to 6419)
  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: "Elife."; 
            date: "2022-09-23"; volume: "11:e79771. doi"; pages: " 
            10.7554/eLife.79771"
COMMENT     Sequence Label: pQE-80L 2xHis-LRRK2 Armadillo 1-552
FEATURES             Location/Qualifiers
     source          1..6419
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(9..23,24..29,30..46,47..53)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    29..45
                     /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)."
     protein_bind    61..77
                     /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)."
     RBS             96..107
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             117..134
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             144..161
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1875..1969
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             2013..2672
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2079..2098)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2737..2823
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2916..3998)
                     /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     3960..3979
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3999..4076)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     complement(4160..4182)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4268..4408
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4323..4342
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4423..4440)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4594..5182)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4674..4693)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5353..6144,6145..6213))
                     /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     5976..5995
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6214..6318)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6386..6404
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgagaaatc ataaaaaatt tatttgcttt gtgagcggat aacaattata atagattcaa
       61 ttgtgagcgg ataacaattt cacacagaat tcattaaaga ggagaaatta actatgcatc
      121 accatcacca tcacagagga tcgcatcacc atcaccatca cggatccatg gctagtggca
      181 gctgtcaggg gtgcgaagag gacgaggaaa ctctgaagaa gttgatagtc aggctgaaca
      241 atgtccagga aggaaaacag atagaaacgc tggtccaaat cctggaggat ctgctggtgt
      301 tcacgtactc cgagcgcgcc tccaagttat ttcaaggcaa aaatatccat gtgcctctgt
      361 tgatcgtctt ggactcctat atgagagtcg cgagtgtgca gcaggtgggt tggtcacttc
      421 tgtgcaaatt aatagaagtc tgtccaggta caatgcaaag cttaatggga ccccaggatg
      481 ttggaaatga ttgggaagtc cttggtgttc accaattgat tcttaaaatg ctaacagttc
      541 ataatgccag tgtaaacttg tcagtgattg gactgaagac cttagatctc ctcctaactt
      601 caggtaaaat caccttgctg atattggatg aagaaagtga tattttcatg ttaatttttg
      661 atgccatgca ctcatttcca gccaatgatg aagtccagaa acttggatgc aaagctttac
      721 atgtgctgtt tgagagagtc tcagaggagc aactgactga atttgttgag aacaaagatt
      781 atatgatatt gttaagtgcg ttaacaaatt ttaaagatga agaggaaatt gtgcttcatg
      841 tgctgcattg tttacattcc ctagcgattc cttgcaataa tgtggaagtc ctcatgagtg
      901 gcaatgtcag gtgttataat attgtggtgg aagctatgaa agcattccct atgagtgaaa
      961 gaattcaaga agtgagttgc tgtttgctcc ataggcttac attaggtaat tttttcaata
     1021 tcctggtatt aaacgaagtc catgagtttg tggtgaaagc tgtgcagcag tacccagaga
     1081 atgcagcatt gcagatctca gcgctcagct gtttggccct cctcactgag actattttct
     1141 taaatcaaga tttagaggaa aagaatgaga atcaagagaa tgatgatgag ggggaagaag
     1201 ataaattgtt ttggctggaa gcctgttaca aagcattaac gtggcataga aagaacaagc
     1261 acgtgcagga ggccgcatgc tgggcactaa ataatctcct tatgtaccaa aacagtttac
     1321 atgagaagat tggagatgaa gatggccatt tcccagctca tagggaagtg atgctctcca
     1381 tgctgatgca ttcttcatca aaggaagttt tccaggcatc tgcgaatgca ttgtcaactc
     1441 tcttagaaca aaatgttaat ttcagaaaaa tactgttatc aaaaggaata cacctgaatg
     1501 ttttggagtt aatgcagaag catatacatt ctcctgaagt ggctgaaagt ggctgtaaaa
     1561 tgctaaatca tctttttgaa ggaagcaaca cttccctgga tataatggca gcagtggtcc
     1621 ccaaaatact aacagttatg aaacgtcatg agacatcatt accagtgcag ctggaggcgc
     1681 ttcgagctat tttacatttt atagtgcctg gcatgccaga agaatccagg gaggatacag
     1741 aatttcatca taagctaaat atggttaaaa aacagtgttt caagaatgat attcacaaac
     1801 tggtcctagc agctttgaac aggtagtcta gaattccctg caggtaatta aataagctta
     1861 attagctgag cttggactcc tgttgataga tccagtaatg acctcagaac tccatctgga
     1921 tttgttcaga acgctcggtt gccgccgggc gttttttatt ggtgagaatc caagctagct
     1981 tggcgagatt ttcaggagct aaggaagcta aaatggagaa aaaaatcact ggatatacca
     2041 ccgttgatat atcccaatgg catcgtaaag aacattttga ggcatttcag tcagttgctc
     2101 aatgtaccta taaccagacc gttcagctgg atattacggc ctttttaaag accgtaaaga
     2161 aaaataagca caagttttat ccggccttta ttcacattct tgcccgcctg atgaatgctc
     2221 atccggaatt tcgtatggca atgaaagacg gtgagctggt gatatgggat agtgttcacc
     2281 cttgttacac cgttttccat gagcaaactg aaacgttttc atcgctctgg agtgaatacc
     2341 acgacgattt ccggcagttt ctacacatat attcgcaaga tgtggcgtgt tacggtgaaa
     2401 acctggccta tttccctaaa gggtttattg agaatatgtt tttcgtctca gccaatccct
     2461 gggtgagttt caccagtttt gatttaaacg tggccaatat ggacaacttc ttcgcccccg
     2521 ttttcaccat gggcaaatat tatacgcaag gcgacaaggt gctgatgccg ctggcgattc
     2581 aggttcatca tgccgtttgt gatggcttcc atgtcggcag aatgcttaat gaattacaac
     2641 agtactgcga tgagtggcag ggcggggcgt aattttttta aggcagttat tggtgccctt
     2701 aaacgcctgg ggtaatgact ctctagcttg aggcatcaaa taaaacgaaa ggctcagtcg
     2761 aaagactggg cctttcgttt tatctgttgt ttgtcggtga acgctctcct gagtaggaca
     2821 aatccgccct ctagattacg tgcagtcgat gataagctgt caaacatgag aattgtgcct
     2881 aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     2941 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     3001 ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc
     3061 accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga
     3121 aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg
     3181 tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt
     3241 gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc
     3301 agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct
     3361 atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc
     3421 gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga
     3481 tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc
     3541 tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg
     3601 gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga
     3661 ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg
     3721 ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc
     3781 agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt
     3841 gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt
     3901 ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg
     3961 gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc
     4021 tcttccgggc gctatcatgc cataccgcga aaggttttgc accattcgat ggtgtcggaa
     4081 tttcgggcag cgttgggtcc tggccacggg tgcgcatgat ctagagctgc ctcgcgcgtt
     4141 tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc
     4201 tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt
     4261 gtcggggcgc agccatgacc cagtcacgta gcgatagcgg agtgtatact ggcttaacta
     4321 tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag
     4381 atgcgtaagg agaaaatacc gcatcaggcg ctcttccgct tcctcgctca ctgactcgct
     4441 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4501 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4561 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4621 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4681 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4741 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4801 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     4861 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     4921 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     4981 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     5041 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5101 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5161 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5221 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5281 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     5341 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     5401 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     5461 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     5521 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5581 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5641 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5701 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5761 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     5821 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     5881 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     5941 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6001 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6061 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6121 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6181 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6241 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6301 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     6361 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtcttcacc
//
