LOCUS       sequence-412900-        6565 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of HA-L2D/V9D/I19D PPM1H.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6565)
  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 6565)
  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: pCMV5D HA-L2D/V9D/I19D PPM1H
FEATURES             Location/Qualifiers
     source          1..6565
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             863..889
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2450..2476
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2517..2534)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2523..2747
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2880..2899)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3031..3078
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             3086..4108
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4238..4359
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4275..4294)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4329..4348
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4408..4424)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4408..4424)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4421..4443)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4432..4448
                     /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)."
     promoter        complement(join(4456..4462,4463..4480,4481..4486))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4501..4522
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4639..4656)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4810..5398)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4890..4909)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5569..6360,6361..6429))
                     /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     6192..6211
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6430..6534)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtccgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta cgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacaccaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta ataaccccgc cccgttgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagaatt
      841 ggccaccgga tctgccgcca tgtacccata cgatgtgcca gattacgccg gatccatgga
      901 cactcgagtg aaatctgccg acgccaattt catgggcggc gacatggctg gcagctcagg
      961 ctccgagcac ggcggcggca gctgcggagg ctcggacctg cccctgcgtt tcccctacgg
     1021 gcggccagag ttcctggggc tgtctcagga cgaggtggag tgcagcgccg accacatcgc
     1081 ccgccccatc ctcatcctca aggagactcg gcggctgccc tgggccactg gctacgcaga
     1141 ggttatcaat gccgggaaga gcacacacaa tgaagaccaa gccagctgtg aggtgctcac
     1201 tgtgaagaag aaggcagggg ccgtgacctc aaccccaaac aggaactcat ccaagagacg
     1261 gtcctccctt cccaatgggg aagggctgca gctgaaggag aactcggaat ccgagggtgt
     1321 ttcctgccac tattggtcgc tgtttgacgg gcacgcgggg tccggggccg cggtggtggc
     1381 gtcacgcctg ctgcagcacc acatcacgga gcagctgcag gacatcgtgg acatcctgaa
     1441 gaactccgcc gtcctgcccc ctacctgcct gggggaggag cctgagaaca cgcccgccaa
     1501 cagccggact ctgacccggg cagcctccct gcgcggaggg gtgggggccc cgggctcccc
     1561 cagcacgccc cccacacgct tctttaccga gaagaagatt ccccatgagt gcctggtcat
     1621 cggagcgctt gaaagtgcat tcaaggaaat ggacctacag atagaacgag agaggagttc
     1681 atataatata tctggtggct gcacggccct cattgtgatt tgccttttgg ggaagctgta
     1741 tgttgcaaat gctggggata gcagggccat aatcatcaga aatggagaaa ttatccccat
     1801 gtcttcagaa tttacccccg agacggagcg ccagcgactt cagtacctgg cattcatgca
     1861 gcctcacttg ctgggaaatg agttcacaca tttggagttt ccaaggagag tacagagaaa
     1921 ggagcttgga aagaagatgc tctacaggga ctttaatatg acaggctggg catacaaaac
     1981 cattgaggat gaggacttga agttccccct tatatatgga gaaggcaaga aggcccgggt
     2041 aatggcaact attggagtga ccaggggact tggggaccat gacctgaagg tgcatgactc
     2101 caacatctac attaaaccat tcctgtcttc agctccagag gtaagaatct acgatctttc
     2161 aaaatatgat catggatcag atgatgtgct gatcttggcc actgatggac tctgggacgt
     2221 tttatcaaat gaagaagtag cagaagcaat cactcagttt cttcctaact gtgatccaga
     2281 tgatcctcac aggtacacac tggcagctca ggacctggtg atgcgtgccc ggggtgtgct
     2341 gaaggacaga ggatggcgga tatctaatga ccgactgggc tcaggagacg acatttctgt
     2401 atatgtcatt cctttaatac atggaaacaa gctgtcatga gcggccgctt acccatacga
     2461 tgtgccagat tacgcctaag tcgagtctag agggcccgtt taaacccgct gatcagcctc
     2521 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     2581 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     2641 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     2701 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     2761 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     2821 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     2881 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     2941 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     3001 acttgattag ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct
     3061 agaaagtata ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa
     3121 gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga
     3181 atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg
     3241 cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc
     3301 gattccggaa gtgcttgaca ttggggagtt cagcgagagc ctgacctatt gcatctcccg
     3361 ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca
     3421 gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt
     3481 cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc
     3541 gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc
     3601 cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca
     3661 cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc
     3721 ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt
     3781 cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca
     3841 tccggagctt gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca
     3901 actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg
     3961 cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag
     4021 cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc
     4081 cagcactcgt ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc
     4141 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     4201 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
     4261 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     4321 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
     4381 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     4441 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     4501 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     4561 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     4621 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     4681 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     4741 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     4801 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     4861 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     4921 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     4981 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     5041 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     5101 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     5161 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     5221 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     5281 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     5341 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     5401 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     5461 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     5521 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     5581 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     5641 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     5701 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     5761 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     5821 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     5881 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     5941 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     6001 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     6061 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     6121 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     6181 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     6241 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     6301 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     6361 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     6421 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     6481 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     6541 tccccgaaaa gtgccacctg acgtc
//
