LOCUS       sequence-366091-        5375 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  2xHis-LRRK2 Armadillo 350-550.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5375)
  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 5375)
  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 350-550
FEATURES             Location/Qualifiers
     source          1..5375
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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    62..78
                     /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             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             118..135
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             145..162
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      832..926
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             970..1629
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1036..1055)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      1694..1780
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1873..2955)
                     /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     2917..2936
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2956..3033)
                     /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(3117..3139)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3225..3365
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3280..3299
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3380..3397)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3551..4139)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3631..3650)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4310..5101,5102..5170))
                     /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     4933..4952
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5171..5275)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5343..5361
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgcat
      121 caccatcacc atcacagagg atcgcatcac catcaccatc acggatccat gctggaagcc
      181 tgttacaaag cattaacgtg gcatagaaag aacaagcacg tgcaggaggc cgcatgctgg
      241 gcactaaata atctccttat gtaccaaaac agtttacatg agaagattgg agatgaagat
      301 ggccatttcc cagctcatag ggaagtgatg ctctccatgc tgatgcattc ttcatcaaag
      361 gaagttttcc aggcatctgc gaatgcattg tcaactctct tagaacaaaa tgttaatttc
      421 agaaaaatac tgttatcaaa aggaatacac ctgaatgttt tggagttaat gcagaagcat
      481 atacattctc ctgaagtggc tgaaagtggc tgtaaaatgc taaatcatct ttttgaagga
      541 agcaacactt ccctggatat aatggcagca gtggtcccca aaatactaac agttatgaaa
      601 cgtcatgaga catcattacc agtgcagctg gaggcgcttc gagctatttt acattttata
      661 gtgcctggca tgccagaaga atccagggag gatacagaat ttcatcataa gctaaatatg
      721 gttaaaaaac agtgtttcaa gaatgatatt cacaaactgg tcctagcagc tttgaacagg
      781 tagtctagaa ttccctgcag gtaattaaat aagcttaatt agctgagctt ggactcctgt
      841 tgatagatcc agtaatgacc tcagaactcc atctggattt gttcagaacg ctcggttgcc
      901 gccgggcgtt ttttattggt gagaatccaa gctagcttgg cgagattttc aggagctaag
      961 gaagctaaaa tggagaaaaa aatcactgga tataccaccg ttgatatatc ccaatggcat
     1021 cgtaaagaac attttgaggc atttcagtca gttgctcaat gtacctataa ccagaccgtt
     1081 cagctggata ttacggcctt tttaaagacc gtaaagaaaa ataagcacaa gttttatccg
     1141 gcctttattc acattcttgc ccgcctgatg aatgctcatc cggaatttcg tatggcaatg
     1201 aaagacggtg agctggtgat atgggatagt gttcaccctt gttacaccgt tttccatgag
     1261 caaactgaaa cgttttcatc gctctggagt gaataccacg acgatttccg gcagtttcta
     1321 cacatatatt cgcaagatgt ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg
     1381 tttattgaga atatgttttt cgtctcagcc aatccctggg tgagtttcac cagttttgat
     1441 ttaaacgtgg ccaatatgga caacttcttc gcccccgttt tcaccatggg caaatattat
     1501 acgcaaggcg acaaggtgct gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat
     1561 ggcttccatg tcggcagaat gcttaatgaa ttacaacagt actgcgatga gtggcagggc
     1621 ggggcgtaat ttttttaagg cagttattgg tgcccttaaa cgcctggggt aatgactctc
     1681 tagcttgagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat
     1741 ctgttgtttg tcggtgaacg ctctcctgag taggacaaat ccgccctcta gattacgtgc
     1801 agtcgatgat aagctgtcaa acatgagaat tgtgcctaat gagtgagcta acttacatta
     1861 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     1921 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct
     1981 tttcaccagt gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg
     2041 cagcaagcgg tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa
     2101 cggcgggata taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc
     2161 accaacgcgc agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt
     2221 ggcaaccagc atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa
     2281 accggacatg gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt
     2341 gagatattta tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc
     2401 taacagcgcg atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc
     2461 gtcttcatgg gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa
     2521 cgccggaaca ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata
     2581 gttaatgatc agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc
     2641 ttcgacgccg cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg
     2701 agatttaatc gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac
     2761 gccaatcagc aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt
     2821 cagctccgcc atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg
     2881 gttcaccacg cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa
     2941 cgttactggt ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat
     3001 accgcgaaag gttttgcacc attcgatggt gtcggaattt cgggcagcgt tgggtcctgg
     3061 ccacgggtgc gcatgatcta gagctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc
     3121 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     3181 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag
     3241 tcacgtagcg atagcggagt gtatactggc ttaactatgc ggcatcagag cagattgtac
     3301 tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     3361 tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3421 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3481 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3541 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3601 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3661 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3721 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     3781 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     3841 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     3901 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     3961 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     4021 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4081 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4141 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     4201 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     4261 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     4321 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     4381 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     4441 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     4501 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     4561 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4621 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4681 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     4741 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     4801 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     4861 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     4921 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     4981 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     5041 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     5101 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     5161 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     5221 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     5281 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     5341 aaaataggcg tatcacgagg ccctttcgtc ttcac
//
