LOCUS       sequence-413046-        7508 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of His SUMO PPM1H.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7508)
  AUTHORS   Wondwossen M Yeshaw , Ayan Adhikari, Claire Y Chiang, Herschel S 
            Dhekne, Paulina S Wawro, Suzanne R Pfeffer
  TITLE     Localization of PPM1H phosphatase tunes Parkinson's disease-linked 
            LRRK2 kinase-mediated Rab GTPase phosphorylation and ciliogenesis
  JOURNAL   Proc Natl Acad Sci U S A. 2023 Oct 31;120(44)
REFERENCE   2  (bases 1 to 7508)
  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: "Proc Natl 
            Acad Sci U S A. 2023 Oct 31;120(44)"
COMMENT     Sequence Label: pET15b+His SUMO PPM1H
FEATURES             Location/Qualifiers
     source          1..7508
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1793..1984
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1978..2000)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2086..2225
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2141..2160
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2240..2257)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2411..2999)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2491..2510)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3170..3961,3962..4030))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3793..3812
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4031..4135)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4203..4221
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(4248..4253,4254..4270,4271..4276)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     terminator      complement(4451..4498)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     4494..4512
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(6410..6427)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             6440..6462
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    6477..6501
                     /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)."
     primer_bind     complement(6501..6520)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6502..6520)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6604..6623)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        6829..6906
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(6907..7508,1..481)
                     /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     complement(6926..6945)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gcataaatat ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag
       61 tgccatgtcc ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc
      121 gatgctggtt gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg
      181 gctgcgcgtt ggtgcggata tctcggtagt gggatacgac gataccgaag acagctcatg
      241 ttatatcccg ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt
      301 ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt
      361 ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct ctccccgcgc
      421 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
      481 agcgcaacgc aattaatgta agttagctca ctcattaggc accgggatct cgaccgatgc
      541 ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg actatcgtcg
      601 ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg gcagcgctct
      661 gggtcatttt cggcgaggac cgctttcgct ggagcgcgac gatgatcggc ctgtcgcttg
      721 cggtattcgg aatcttgcac gccctcgctc aagccttcgt cactggtccc gccaccaaac
      781 gtttcggcga gaagcaggcc attatcgccg gcatggcggc cgacgcgctg ggctacgtct
      841 tgctggcgtt cgcgacgcga ggctggatgg ccttccccat tatgattctt ctcgcttccg
      901 gcggcatcgg gatgcccgcg ttgcaggcca tgctgtccag gcaggtagat gacgaccatc
      961 agggacagct tcaaggatcg ctcgcggctc ttaccagcct aacttcgatc attggaccgc
     1021 tgatcgtcac ggcgatttat gccgcctcgg cgagcacatg gaacgggttg gcatggattg
     1081 taggcgccgc cctatacctt gtctgcctcc ccgcgttgcg tcgcggtgca tggagccggg
     1141 ccacctcgac ctgaatggaa gccggcggca cctcgctaac ggattcacca ctccaagaat
     1201 tggagccaat caattcttgc ggagaactgt gaatgcgcaa accaaccctt ggcagaacat
     1261 atccatcgcg tccgccatct ccagcagccg cacgcggcgc atctcgggca gcgttgggtc
     1321 ctggccacgg gtgcgcatga tcgtgctcct gtcgttgagg acccggctag gctggcgggg
     1381 ttgccttact ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa gcgactgctg
     1441 ctgcaaaacg tctgcgacct gagcaacaac atgaatggtc ttcggtttac gtgtttcgta
     1501 aagtctggaa acgcggaagt cagcgccctg caccattatg ttccggatct gcatcgcagg
     1561 atgctgctgg ctaccctgtg gaacacctac atctgtatta acgaagcgct ggcattgacc
     1621 ctgagtgatt tttctctggt cccgccgcat ccataccgcc agttgtttac cctcacaacg
     1681 ttccagtaac cgggcatgtt catcatcagt aacccgtatc gtgagcatcc tctctcgttt
     1741 catcggtatc attaccccca tgaacagaaa tcccccttac acggaggcat cagtgaccaa
     1801 acaggaaaaa accgccctta acatggcccg ctttatcaga agccagacat taacgcttct
     1861 ggagaaactc aacgagctgg acgcggatga acaggcagac atctgtgaat cgcttcacga
     1921 ccacgctgat gagctttacc gcagctgcct cgcgcgtttc ggtgatgacg gtgaaaacct
     1981 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag
     2041 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca
     2101 gtcacgtagc gatagcggag tgtatactgg cttaactatg cggcatcaga gcagattgta
     2161 ctgagagtgc accattgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     2221 tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     2281 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     2341 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     2401 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     2461 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     2521 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     2581 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     2641 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     2701 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     2761 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     2821 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     2881 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     2941 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     3001 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     3061 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     3121 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     3181 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     3241 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     3301 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     3361 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     3421 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     3481 ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     3541 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     3601 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     3661 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     3721 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     3781 cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     3841 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     3901 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     3961 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     4021 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     4081 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     4141 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     4201 aaaataggcg tatcacgagg ccctttcgtc ttcaagaagt ctcatgtttg acagcttatc
     4261 atcgataagc tttaatgcgg tagtttatca cagttaaatt gctaacgcag tcaggcaccg
     4321 tgtatgaaat ctaacaatgc gctcatcgtc atcctcggca ccgtcaccct ggatgctgta
     4381 ggcataggct tggttatgcc ggtactgccg ggcctcttgc gggatatccg gatatagttc
     4441 ctcctttcag caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt tatgctagtt
     4501 attgctcagc ggtggcagca gccaactcag cttcctttcg ggctttgtta gcagccggat
     4561 ctttacgcgg ccgtcatcat gacagcttgt ttccatgtat taaaggaatg acatatacag
     4621 aaatgtcgtc tcctgagccc agtcggtcat tagatatccg ccatcctctg tccttcagca
     4681 caccccgggc acgcatcacc aggtcctgag ctgccagtgt gtacctgtga ggatcatctg
     4741 gatcacagtt aggaagaaac tgagtgattg cttctgctac ttcttcattt gataaaacgt
     4801 cccagagtcc atcagtggcc aagatcagca catcatctga tccatgatca tattttgaaa
     4861 gatcgtagat tcttacctct ggagctgaag acaggaatgg tttaatgtag atgttggagt
     4921 catgcacctt caggtcatgg tccccaagtc ccctggtcac tccaatagtt gccattaccc
     4981 gggccttctt gccttctcca tatataaggg ggaacttcaa gtcctcatcc tcaatggttt
     5041 tgtatgccca gcctgtcata ttaaagtccc tgtagagcat cttctttcca agctcctttc
     5101 tctgtactct ccttggaaac tccaaatgtg tgaactcatt tcccagcaag tgaggctgca
     5161 tgaatgccag gtactgaagt cgctggcgct ccgtctcggg ggtaaattct gaagacatgg
     5221 ggataatttc tccatttctg atgattatgg ccctgctatc cccagcattt gcaacataca
     5281 gcttccccaa aaggcaaatc acaatgaggg ccgtgcagcc accagatata ttatatgaac
     5341 tcctctctcg ttctatctgt aggtccattt ccttgaatgc actttcaagc gctccgatga
     5401 ccaggcactc atggggaatc ttcttctcgg taaagaagcg tgtggggggc gtgctggggg
     5461 agcccggggc ccccacccct ccgcgcaggg aggctgcccg ggtcagagtc cggctgttgg
     5521 cgggcgtgtt ctcaggctcc tcccccaggc aggtaggggg caggacggcg gagttcttca
     5581 ggatgtccac gatgtcctgc agctgctccg tgatgtggtg ctgcagcagg cgtgacgcca
     5641 ccaccgcggc cccggacccc gcgtgcccgt caaacagcga ccaatagtgg caggaaacac
     5701 cctcggattc cgagttctcc ttcagctgca gcccttcccc attgggaagg gaggaccgtc
     5761 tcttggatga gttcctgttt ggggttgagg tcacggcccc tgccttcttc ttcacagtga
     5821 gcacctcaca gctggcttgg tcttcattgt gtgtgctctt cccggcattg ataacctctg
     5881 cgtagccagt ggcccagggc agccgccgag tctccttgag gatgaggatg gggcgggcga
     5941 tgtggtcggc gctgcactcc acctcgtcct gagacagccc caggaactct ggccgcccgt
     6001 aggggaaacg caggggcagg tccgagcctc cgcagctgcc gccgccgtgc tcggagcctg
     6061 agctgccagc catgatgccg cccatgaaat tggccacggc agatttcact cgagtgagca
     6121 ttccccccgt ttgttcctga taaacttcaa tcacatcttc ttcctccatt cccagttctt
     6181 ttggagtatg attatcagca attctctgac cctcaaagag aaacctgagt gagttcattg
     6241 gaacaccctg tctttgacag tatgattctt tgagtttctt gagatgtgtt gtcattttca
     6301 ctttgaagtg aatctcactg ctatcctgtc caatgacttt gagtttaata tattcacctt
     6361 ccttcttatc ccccaagtcc tcagttgaag gttttgcctc ctggtcagaa tggtgatggt
     6421 gatgatgacc catggtatat ctccttctta aagttaaaca aaattatttc tagaggggaa
     6481 ttgttatccg ctcacaattc ccctatagtg agtcgtatta atttcgcggg atcgagatct
     6541 cgatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt gcggttgctg
     6601 gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc gggctcatga
     6661 gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg ttgggcgcca
     6721 tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac ctactactgg
     6781 gctgcttcct aatgcaggag tcgcataagg gagagcgtcg agatcccgga caccatcgaa
     6841 tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg
     6901 gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag
     6961 accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg
     7021 gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc
     7081 aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa
     7141 attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg
     7201 gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc
     7261 gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct
     7321 gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt
     7381 attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt
     7441 caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg
     7501 gctggctg
//
