LOCUS       sequence-232194-        6625 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian overexpression of StrepII-tagged full length PPME1 with 
            mutation in the methylesterase active center.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6625)
  TITLE     Westermarck PPME1 expression plasmids
REFERENCE   2  (bases 1 to 6625)
  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"
COMMENT     Sequence Label: PME-1 S156A (inakt.mut) pcDNA-StrepIII
FEATURES             Location/Qualifiers
     source          1..6625
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1863..1886
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1923..1946
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(1982..1999)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1988..2212
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2258..2686
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2345..2364)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2555..2576
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2695..2715)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2700..3029
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2880..3015
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2942..2961
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3096..3890
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3150..3169)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3760..3779
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4064..4185
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4101..4120)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4155..4174
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4234..4250)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4234..4250)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4247..4269)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4258..4274
                     /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(4282..4288,4289..4306,4307..4312))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4327..4348
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4465..4482)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4636..5224)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4716..4735)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5395..6186,6187..6255))
                     /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     6018..6037
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6256..6360)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6435..6454)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacgggccc tctagactcg agatgtcggc cctcgaaaag agcatgcacc
      721 tcggccgcct tccctctcgc ccacctctac ccggcagcgg gggcagtcag agcggagcca
      781 agatgcgaat gggccctgga agaaagcggg acttttcccc tgttccttgg agtcagtatt
      841 ttgagtccat ggaagatgta gaagtagaga atgaaactgg caaggatact tttcgagtct
      901 acaagagtgg ttcagagggt ccagtcctgc tccttctgca tggaggaggt cattctgccc
      961 tttcttgggc tgtgttcacg gcagcgatta ttagtagagt tcagtgtagg attgtagctt
     1021 tggatctgcg aagtcatggt gaaacaaagg tcaagaatcc tgaagatctg tctgcagaaa
     1081 caatggcaaa agacgttggc aatgtggttg aagccatgta tggggacctt cctcctccaa
     1141 ttatgctgat tggacatgcc atgggtggtg ctattgcagt ccacacagca tcatccaacc
     1201 tggtaccaag cctcttgggt ctgtgcatga ttgatgttgt agaaggtaca gctatggatg
     1261 cacttaatag catgcagaat ttcttacggg gtcgtcctaa aaccttcaag tctctggaga
     1321 atgctattga atggagtgtg aagagtggcc agattcgaaa tctggagtct gcccgtgtct
     1381 caatggttgg ccaagtcaaa cagtgtgaag gaattacaag tccagaaggc tcaaaatcta
     1441 tagtggaagg aatcatagag gaagaagaag aagatgagga aggaagtgag tctataagca
     1501 agaggaaaaa ggaagatgac atggagacca agaaagacca tccatacacc tggagaattg
     1561 aactggcaaa aacagaaaaa tactgggacg gctggttccg aggcttatcc aatctctttc
     1621 ttagttgtcc cattcctaaa ttgctgctct tggctggtgt tgatagattg gataaagatc
     1681 tgaccattgg ccagatgcaa gggaagttcc agatgcaggt cctaccccag tgtggccatg
     1741 cagtccatga ggatgcccct gacaaggtag ctgaagctgt tgccactttc ctgatccggc
     1801 acaggtttgc agaacccatc ggtggattcc agtgtgtgtt tcctggctgt ggatccgcta
     1861 gctggagcca cccgcagttc gagaaaggtg gaggttccgg aggtggatcg ggaggtggat
     1921 cgtggagcca cccgcagttc gaaaaaggcg cctaaaagct taagtttaaa ccgctgatca
     1981 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
     2041 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg
     2101 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg
     2161 gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag
     2221 gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta
     2281 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     2341 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     2401 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     2461 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     2521 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     2581 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     2641 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg
     2701 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     2761 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     2821 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     2881 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     2941 ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag
     3001 gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg
     3061 gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     3121 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     3181 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     3241 tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt
     3301 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     3361 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     3421 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     3481 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     3541 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     3601 aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg
     3661 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     3721 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     3781 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     3841 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     3901 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     3961 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     4021 cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc
     4081 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     4141 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc
     4201 gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     4261 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     4321 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4381 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4441 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4501 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4561 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     4621 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     4681 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     4741 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     4801 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     4861 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     4921 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     4981 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     5041 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     5101 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     5161 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5221 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5281 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5341 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5401 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5461 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     5521 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     5581 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     5641 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     5701 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     5761 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     5821 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     5881 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     5941 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     6001 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     6061 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     6121 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     6181 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6241 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6301 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6361 cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta
     6421 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
     6481 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa
     6541 ggcttgaccg acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc
     6601 gatgtacggg ccagatatac gcgtt
//
