LOCUS       sequence-474744-        6216 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses PPM1H in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6216)
  AUTHORS   Wang X, Xu P, Bentley-DeSousa A, Hancock-Cerutti W, Cai S, Johnson 
            BT, Tonelli F, Shao L, Talaia G, Alessi DR, Ferguson SM, De Camilli 
            P
  TITLE     The bridge-like lipid transport protein VPS13C/PARK23 mediates 
            ER-lysosome contacts following lysosome damage.
  JOURNAL   Nat Cell Biol. 2025 May;27(5):776-789. doi: 
            10.1038/s41556-025-01653-6. Epub 2025 Apr 10.
  PUBMED    40211074
REFERENCE   2  (bases 1 to 6216)
  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"; doi: 
            "10.1038/s41556-025-01653-6"; journalName: "Nat Cell Biol"; date: 
            "2025-05"; volume: "27"; issue: "5"; pages: "776-789"
COMMENT     Sequence Label: mScarlet-PPM1H
FEATURES             Location/Qualifiers
     source          1..6216
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             613..1308
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(746..764)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1014..1033
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    3004..3125
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3041..3060)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3095..3114
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3132..3587)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3269..3290)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3481..3500
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3614..3718
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3720..4077
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3720..3911
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3743..3763)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3928..4063
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4112..4906
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4112..4906
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4166..4185)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4776..4795
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5094..5113)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5138..5185
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5514..6102
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6003..6022
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gcagtgatca aggagttcat gcggttcaag
      661 gtgcacatgg agggctccat gaacggccac gagttcgaga tcgagggcga gggcgagggc
      721 cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc
      781 ttctcctggg acatcctgtc ccctcagttc atgtacggct ccagggcctt catcaagcac
      841 cccgccgaca tccccgacta ctataagcag tccttccccg agggcttcaa gtgggagcgc
      901 gtgatgaact tcgaggacgg cggcgccgtg accgtgaccc aggacacctc cctggaggac
      961 ggcaccctga tctacaaggt gaagctccgc ggcaccaact tccctcctga cggccccgta
     1021 atgcagaaga agacaatggg ctgggaagcg tccaccgagc ggttgtaccc cgaggacggc
     1081 gtgctgaagg gcgacattaa gatggccctg cgcctgaagg acggcggccg ctacctggcg
     1141 gacttcaaga ccacctacaa ggccaagaag cccgtgcaga tgcccggcgc ctacaacgtc
     1201 gaccgcaagt tggacatcac ctcccacaac gaggactaca ccgtggtgga acagtacgaa
     1261 cgctccgagg gccgccactc caccggcggc atggacgagc tgtacaagtc cggactcaga
     1321 tctcgagcta tgctcactcg agtgaaatct gccgtggcca atttcatggg cggcatcatg
     1381 gctggcagct caggctccga gcacggcggc ggcagctgcg gaggctcgga cctgcccctg
     1441 cgtttcccct acgggcggcc agagttcctg gggctgtctc aggacgaggt ggagtgcagc
     1501 gccgaccaca tcgcccgccc catcctcatc ctcaaggaga ctcggcggct gccctgggcc
     1561 actggctacg cagaggttat caatgccggg aagagcacac acaatgaaga ccaagccagc
     1621 tgtgaggtgc tcactgtgaa gaagaaggca ggggccgtga cctcaacccc aaacaggaac
     1681 tcatccaaga gacggtcctc ccttcccaat ggggaagggc tgcagctgaa ggagaactcg
     1741 gaatccgagg gtgtttcctg ccactattgg tcgctgtttg acgggcacgc ggggtccggg
     1801 gccgcggtgg tggcgtcacg cctgctgcag caccacatca cggagcagct gcaggacatc
     1861 gtggacatcc tgaagaactc cgccgtcctg ccccctacct gcctggggga ggagcctgag
     1921 aacacgcccg ccaacagccg gactctgacc cgggcagcct ccctgcgcgg aggggtgggg
     1981 gccccgggct cccccagcac gccccccaca cgcttcttta ccgagaagaa gattccccat
     2041 gagtgcctgg tcatcggagc gcttgaaagt gcattcaagg aaatggacct acagatagaa
     2101 cgagagagga gttcatataa tatatctggt ggctgcacgg ccctcattgt gatttgcctt
     2161 ttggggaagc tgtatgttgc aaatgctggg gatagcaggg ccataatcat cagaaatgga
     2221 gaaattatcc ccatgtcttc agaatttacc cccgagacgg agcgccagcg acttcagtac
     2281 ctggcattca tgcagcctca cttgctggga aatgagttca cacatttgga gtttccaagg
     2341 agagtacaga gaaaggagct tggaaagaag atgctctaca gggactttaa tatgacaggc
     2401 tgggcataca aaaccattga ggatgaggac ttgaagttcc cccttatata tggagaaggc
     2461 aagaaggccc gggtaatggc aactattgga gtgaccaggg gacttgggga ccatgacctg
     2521 aaggtgcatg actccaacat ctacattaaa ccattcctgt cttcagctcc agaggtaaga
     2581 atctacgatc tttcaaaata tgatcatgga tcagatgatg tgctgatctt ggccactgat
     2641 ggactctggg acgttttatc aaatgaagaa gtagcagaag caatcactca gtttcttcct
     2701 aactgtgatc cagatgatcc tcacaggtac acactggcag ctcaggacct ggtgatgcgt
     2761 gcccggggtg tgctgaagga cagaggatgg cggatatcta atgaccgact gggctcagga
     2821 gacgacattt ctgtatatgt cattccttta atacatggaa acaagctgtc atgaggatcc
     2881 accggatcta gataactgat cataatcagc cataccacat ttgtagaggt tttacttgct
     2941 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt
     3001 gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     3061 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     3121 tcttaacgcg taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     3181 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     3241 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     3301 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     3361 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     3421 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     3481 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     3541 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt
     3601 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     3661 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtcc
     3721 tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     3781 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     3841 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     3901 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     3961 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     4021 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat
     4081 cgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     4141 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     4201 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     4261 gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct
     4321 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     4381 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     4441 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     4501 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     4561 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     4621 gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     4681 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     4741 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     4801 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     4861 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     4921 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc
     4981 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     5041 cagcgcgggg atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca
     5101 cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa
     5161 acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     5221 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     5281 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     5341 cctgccatag cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa
     5401 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     5461 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     5521 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     5581 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5641 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     5701 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     5761 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     5821 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     5881 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     5941 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     6001 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     6061 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     6121 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     6181 cctgattctg tggataaccg tattaccgcc atgcat
//
