LOCUS       sequence-486752-        8105 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses His SUMO mNeon (inserted in loop1) Delta-37 PPM1H in 
            bacterial cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8105)
  AUTHORS   Adhikari A, Tripathi A, Chiang CY, Sherpa P, Pfeffer SR
  TITLE     Allosteric regulation of the Golgi-localized PPM1H phosphatase by 
            Rab GTPases modulates LRRK2 substrate dephosphorylation in 
            Parkinson's disease.
  JOURNAL   J Biol Chem. 2025 Sep 3:110679. doi: 10.1016/j.jbc.2025.110679.
  PUBMED    40912655
REFERENCE   2  (bases 1 to 8105)
  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. 2025 Sep 3:110679. doi: 10.1016/j.jbc.2025.110679."
COMMENT     Sequence Label: pET15b His SUMO mNeon  Delta-37 PPM1H
FEATURES             Location/Qualifiers
     source          1..8105
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(18..36)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        104..208
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(209..277,278..1069)
                     /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     complement(427..446)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1240..1828
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1729..1748
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1982..1999
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2014..2153
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2079..2098)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2239..2261
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2255..2446)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3758..4840)
                     /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     4802..4821
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4841..4918)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5124..5143
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5227..5246
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5227..5245
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5246..5270
                     /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             5285..5307
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5320..5337
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(7832..7850)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      7846..7893
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(8068..8073,8074..8090,8091..8096))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
ORIGIN
        1 ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
       61 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
      121 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
      181 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
      241 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
      301 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
      361 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
      421 agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg
      481 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
      541 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
      601 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
      661 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
      721 accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact
      781 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
      841 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
      901 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
      961 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     1021 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     1081 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     1141 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     1201 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     1261 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     1321 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     1381 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     1441 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     1501 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     1561 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     1621 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     1681 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     1741 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     1801 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     1861 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     1921 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     1981 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     2041 tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca
     2101 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     2161 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     2221 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     2281 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
     2341 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     2401 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     2461 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     2521 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     2581 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     2641 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     2701 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgt aaaccgaaga
     2761 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     2821 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     2881 tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgagatgc
     2941 gccgcgtgcg gctgctggag atggcggacg cgatggatat gttctgccaa gggttggttt
     3001 gcgcattcac agttctccgc aagaattgat tggctccaat tcttggagtg gtgaatccgt
     3061 tagcgaggtg ccgccggctt ccattcaggt cgaggtggcc cggctccatg caccgcgacg
     3121 caacgcgggg aggcagacaa ggtatagggc ggcgcctaca atccatgcca acccgttcca
     3181 tgtgctcgcc gaggcggcat aaatcgccgt gacgatcagc ggtccaatga tcgaagttag
     3241 gctggtaaga gccgcgagcg atccttgaag ctgtccctga tggtcgtcat ctacctgcct
     3301 ggacagcatg gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa gaatcataat
     3361 ggggaaggcc atccagcctc gcgtcgcgaa cgccagcaag acgtagccca gcgcgtcggc
     3421 cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac
     3481 gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt
     3541 cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc
     3601 tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc
     3661 ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc
     3721 ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     3781 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     3841 tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct
     3901 tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc
     3961 gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt
     4021 cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca
     4081 ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat
     4141 tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg
     4201 ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg
     4261 ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca
     4321 gatgctccac gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg
     4381 tctggtcaga gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa
     4441 tggcatcctg gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa
     4501 gattgtgcac cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca
     4561 cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt
     4621 gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt
     4681 gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg
     4741 ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac
     4801 cggcatactc tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac
     4861 tctcttccgg gcgctatcat gccataccgc gaaaggtttt gcgccattcg atggtgtccg
     4921 ggatctcgac gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg
     4981 aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc gcccaacagt
     5041 cccccggcca cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag
     5101 tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct
     5161 gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg
     5221 cgaaattaat acgactcact ataggggaat tgtgagcgga taacaattcc cctctagaaa
     5281 taattttgtt taactttaag aaggagatat accatgggtc atcatcacca tcaccattct
     5341 gaccaggagg caaaaccttc aactgaggac ttgggggata agaaggaagg tgaatatatt
     5401 aaactcaaag tcattggaca ggatagcagt gagattcact tcaaagtgaa aatgacaaca
     5461 catctcaaga aactcaaaga atcatactgt caaagacagg gtgttccaat gaactcactc
     5521 aggtttctct ttgagggtca gagaattgct gataatcata ctccaaaaga actgggaatg
     5581 gaggaagaag atgtgattga agtttatcag gaacaaacgg ggggaatgcg tttcccctac
     5641 gggcggccag agttcctggg gctgtctcag gacgaggtgg agtgcagcgc cgaccacatc
     5701 gcccgcccca tcctcatcct caaggagact cggcggctgc cctgggccac tggctacgca
     5761 gaggttatca atgccgggaa gagcacacac aatgaagacc aagccagctg tgaggtgctc
     5821 actgtgaaga agaaggcagg ggccgtgacc tcaaccccaa acaggaactc atccaagaga
     5881 cggtcctcca tggtctccaa aggcgaagag gataacatgg caagtctgcc cgcaacccat
     5941 gagcttcaca ttttcgggtc cataaatgga gtagacttcg atatggtggg tcaaggcaca
     6001 ggtaatccaa atgatgggta cgaggagttg aacctcaaaa gtaccaaagg tgaccttcaa
     6061 tttagtccat ggatcttggt gccgcacata ggatacggat ttcatcaata cctgccgtac
     6121 cccgacggaa tgagtccatt ccaggcagct atggttgatg gatctggcta ccaggttcac
     6181 cgcacaatgc aattcgagga tggagcctca cttacggtga actatcgata tacttatgaa
     6241 ggctcacaca taaaagggga agcccaggta aagggcacag gttttccagc cgatggtccg
     6301 gttatgacaa actctctgac tgctgcagat tggtgtagga gtaaaaagac atacccgaat
     6361 gacaaaacca tcatctcaac tttcaagtgg tcatatacga cgggcaacgg aaagcggtat
     6421 agatccacag ctaggacgac gtacacgttt gctaagccga tggcagcgaa ttatcttaaa
     6481 aatcagccga tgtacgtttt tcggaaaacg gaactgaaac actcaaaaac tgaacttaac
     6541 tttaaggagt ggcagaaagc ctttaccgac gtgatgggaa tggacgaact ctataagccc
     6601 aatggggaag ggctgcagct gaaggagaac tcggaatccg agggtgtttc ctgccactat
     6661 tggtcgctgt ttgacgggca cgcggggtcc ggggccgcgg tggtggcgtc acgcctgctg
     6721 cagcaccaca tcacggagca gctgcaggac atcgtggaca tcctgaagaa ctccgccgtc
     6781 ctgcccccta cctgcctggg ggaggagcct gagaacacgc ccgccaacag ccggactctg
     6841 acccgggcag cctccctgcg cggaggggtg ggggccccgg gctcccccag cacgcccccc
     6901 acacgcttct ttaccgagaa gaagattccc catgagtgcc tggtcatcgg agcgcttgaa
     6961 agtgcattca aggaaatgga cctacagata gaacgagaga ggagttcata taatatatct
     7021 ggtggctgca cggccctcat tgtgatttgc cttttgggga agctgtatgt tgcaaatgct
     7081 ggggatagca gggccataat catcagaaat ggagaaatta tccccatgtc ttcagaattt
     7141 acccccgaga cggagcgcca gcgacttcag tacctggcat tcatgcagcc tcacttgctg
     7201 ggaaatgagt tcacacattt ggagtttcca aggagagtac agagaaagga gcttggaaag
     7261 aagatgctct acagggactt taatatgaca ggctgggcat acaaaaccat tgaggatgag
     7321 gacttgaagt tcccccttat atatggagaa ggcaagaagg cccgggtaat ggcaactatt
     7381 ggagtgacca ggggacttgg ggaccatgac ctgaaggtgc atgactccaa catctacatt
     7441 aaaccattcc tgtcttcagc tccagaggta agaatctacg atctttcaaa atatgatcat
     7501 ggatcagatg atgtgctgat cttggccact gatggactct gggacgtttt atcaaatgaa
     7561 gaagtagcag aagcaatcac tcagtttctt cctaactgtg atccagatga tcctcacagg
     7621 tacacactgg cagctcagga cctggtgatg cgtgcccggg gtgtgctgaa ggacagagga
     7681 tggcggatat ctaatgaccg actgggctca ggagacgaca tttctgtata tgtcattcct
     7741 ttaatacatg gaaacaagct gtcatgatga cggccgcgta aagatccggc tgctaacaaa
     7801 gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt
     7861 ggggcctcta aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggatat
     7921 cccgcaagag gcccggcagt accggcataa ccaagcctat gcctacagca tccagggtga
     7981 cggtgccgag gatgacgatg agcgcattgt tagatttcat acacggtgcc tgactgcgtt
     8041 agcaatttaa ctgtgataaa ctaccgcatt aaagcttatc gatgataagc tgtcaaacat
     8101 gagac
//
