LOCUS       sequence-139990-        8648 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express human Plekhm1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8648)
  AUTHORS   Witwicka H, Jia H, Kutikov A, Reyes-Gutierrez P, Li X, Odgren PR
  TITLE     TRAFD1 (FLN29) Interacts with Plekhm1 and Regulates Osteoclast 
            Acidification and Resorption.
  JOURNAL   PLoS One. 2015 May 19;10(5):e0127537. doi: 
            10.1371/journal.pone.0127537. eCollection 2015.
  PUBMED    25992615
REFERENCE   2  (bases 1 to 8648)
  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: "PLoS One.";
            date: "2015-05-19"; volume: "10(5):e0127537. doi"; pages: " 
            10.1371/journal.pone.0127537. eCollection 2015"
COMMENT     Sequence Label: hPlekhm1_pIRES puro Glue
FEATURES             Location/Qualifiers
     source          1..8648
                     /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..612
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        613..816
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     767..787
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     861..880
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        861..879
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      921..930
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             927..1040
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     CDS             1059..1079
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1086..1112
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             1086..1112
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1113..1190
                     /codon_start=1
                     /product="calmodulin-binding peptide"
                     /label=CBP
                     /note="derived from skeletal muscle myosin light chain 
                     kinase; binds calmodulin with nanomolar affinity in the 
                     presence of calcium"
                     /translation="KRRWKKNFIAVSAANRFKKISSSGAL"
     intron          4507..4736
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     4719..4738
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(4794..5368,5369..5371,5372..5380)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4961..4978)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     5188..5207
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             5400..5999
                     /codon_start=1
                     /gene="pac from Streptomyces"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVECPKDRATWCMTRKPGA"
     primer_bind     complement(5400..5419)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5896..5916
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(6135..6152)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6141..6365
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     6412..6431
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6491..6507)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6491..6507)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6504..6526)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6515..6531
                     /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(6539..6545,6546..6563,6564..6569))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6584..6605
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6722..6739)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6893..7481)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6973..6992)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7652..8443,8444..8512))
                     /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     8275..8294
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8513..8617)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct 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 tagtatcaat tacggggtca ttagttcata gcccatatat
      301 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccacgacccc
      361 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
      421 tgacgtcaat gggtggacta tttacggtaa actgcccact tggcagtaca tcaagtgtat
      481 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
      541 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
      601 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
      661 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
      721 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
      781 aggcgtgtac ggtgggaggt ctatataagc agagctctct ggctaactag agaacccact
      841 gcttactggc ttatcgaaat taatacgact cactataggg agacccaagc ttggtaccga
      901 gctcggatcg atttaattaa gccaccatgg acgagaagac caccggctgg cggggcggcc
      961 acgtggtgga gggcctggcc ggcgagctgg agcagctgcg ggcccggctg gagcaccacc
     1021 cccagggcca gcgggagccc ggcggaagcc ccggtggcga gaacctgtac ttccagggcg
     1081 caagctaccc ctacgacgtg cccgactacg ccaagcggcg gtggaagaag aacttcatcg
     1141 ccgtgagcgc cgccaaccgg ttcaagaaga tcagcagcag cggcgccctg attgatgggg
     1201 gatctggccc cggcggcggc gcgcctgtac agatatcaat gctttcagtg gtggagaatg
     1261 gactggaccc ccaggctgcc atcccggtca tcaagaagaa gctggtggga tccgtgaagg
     1321 ccttgcagaa gcagtacgtg tccctggaca cggtggtcac tagtgaagac ggagatgcca
     1381 acaccatgtg cagcgccctg gaggccgtat ttatccatgg cctgcacgcc aagcacatcc
     1441 gagctgaggc cggaggaaaa aggaagaaaa gtgcccacca gaagcctctg ccccagcctg
     1501 tcttctggcc cctcctgaaa gctgtcaccc acaaacacat catctcagag ttggagcacc
     1561 tgacgtttgt caacacggat gtgggccgct gccgggcatg gctgcggctg gccctgaacg
     1621 atggcctgat ggagtgctac ctgaagctgc tgctgcagga gcaggcccgc ttgcatgagt
     1681 actaccagcc caccgccctg ctccgggatg ctgaggaggg cgagttcctc cttagcttcc
     1741 tgcagggcct cacgtccttg tccttcgaac tctcctacaa gtctgccatc ttaaatgagt
     1801 ggacgctcac cccattggcc ctgtctgggc tttgcccgct ttctgagctg gaccctctct
     1861 ctacctctgg tgcagaacta cagcggaagg aatctctgga ttccatttcc cattcttcag
     1921 gctctgaaga catcgaagtc catcactcgg gccataagat acggaggaac cagaagctga
     1981 ctgcctcctc cctcagcctg gacacggcca gttcatccca gctgtcctgc agcctaaact
     2041 ctgatagctg cttactccaa gagaatggct ccaagagtcc agaccattgc gaggagccca
     2101 tgtcctgtga ctcagacctg ggcacagcaa atgctgagga ctcagaccgg tctctgcaag
     2161 aggtattgtt ggaattcagc aaagcccagg taaactctgt gccaaccaac ggactgagcc
     2221 aagaaacaga gatccccaca ccacaggcct cgctctccct ccatggcctc aacaccagca
     2281 catacctgca ctgtgaggca cctgcagagc cccttcctgc ccaggcagcc tctggaactc
     2341 aagatggtgt ccacgtgcag gagccgcgtc cccaggcgcc cagccccctg gacttacagc
     2401 agcctgtaga gagcacctca ggccagcagc cttctagtac tgtcagcgag acagccagag
     2461 aagtgggcca agggaatggc ctgcagaagg cccaggctca tgacggagct ggtctgaagc
     2521 tggtagtttc ctcacccacc agtccgaaaa acaagagctg gatctcagag gatgacttct
     2581 accggccttc ccgggagcaa cccctggaga gtgcttcaga ccacccaata gcttcttaca
     2641 gggggactcc agggtcaagg cctggtctcc acaggcattt ttctcaagaa ccaagaaaaa
     2701 actgctccct gggggcgtta gaccaagcgt gtgtaccttc cccaggaaga aggcaagccc
     2761 aggcagcccc atcccagggg cataagagct tccgggtggt acaccggaga cagatgggac
     2821 tgtccaaccc attccggggt ctcatgaagc tgggcaccgt ggagcggcgg ggggcaatgg
     2881 gcatctggaa ggagctcttc tgcgagctct ccccgctgga gttccgcctc tacctgagca
     2941 acgaggagca cacctgtgtg gagaactgct cgctgcttcg ctgtgagtct gtggggccag
     3001 cccatagtga tgggcgcttt gagctggtct tctctggcaa gaagctggcc ctgcgcgcct
     3061 cctcccagga cgaagctgag gactggctgg accgggtgcg ggaggccctg cagaaggtcc
     3121 ggcctcagca ggaggatgag tgggtgaacg tgcagtaccc agaccagcct gaggaacccc
     3181 ccgaggcgcc ccagggctgc ctctctccct cagacctgct ctcggagccc gcggccctcc
     3241 agggcacaca gtttgactgg tcgtccgccc aggttccaga gccagatgcc atcaaggagt
     3301 ccctgctgta cttgtacatg gacaggacct ggatgcccta tatattttct ctgtccttgg
     3361 aggctctgaa atgtttccgc atcaggaaca atgagaagat gctgagtgac agccacggcg
     3421 tggagaccat ccgggacatc ctgccagaca ccagccttgg gggcccatcc ttcttcaaaa
     3481 tcatcacggc caaggctgtc ctgaagctgc aggccggaaa cgccgaggaa gccgccctgt
     3541 ggagggatct ggtccgcaaa gtcctggcat cctacttgga gacagccgag gaggcggtga
     3601 ccctgggcgg gagcctggat gaaaactgtc aggaggtgct gaaatttgcc acccgggaga
     3661 atggcttcct gctgcagtac ctggtggcta tccccatgga gaaaggcctt gactcccaag
     3721 gctgcttctg cgcaggctgc tcccggcaga tcggcttctc ctttgtacga cccaagctct
     3781 gtgccttctc tggcctctat tactgtgaca tctgccacca agacgatgcc tcagtgattc
     3841 cggccaggat catccacaac tgggacctca ccaagcgccc gatctgcagg caggccctga
     3901 agtttctgac acagatccgg gcccagcccc tcatcaacct gcagatggtg aacgcgtctc
     3961 tgtacgagca tgtggagcgg atgcacctca ttgggaggag acgggagcag ctgaagctcc
     4021 tgggggatta cctgggcctg tgccggagtg gcgccctgaa ggagctcagc aagaggctca
     4081 accacaggaa ttatctcttg gaatctccgc ataggttcag tgttgctgac ctccaacaga
     4141 tcgcagacgg ggtgtatgaa ggattcctca aggccctgat tgaatttgcc tcccagcatg
     4201 tctaccactg cgacctgtgc acccagcgcg gcttcatctg ccagatctgc cagcaccacg
     4261 acatcatctt cccctttgag tttgacacca cagtcaggtg tgccgagtgc aagaccgtct
     4321 tccaccagag ctgccaggct gtggtgaaga agggctgccc ccgctgtgcc cgccggcgca
     4381 agtaccagga acagaacatt ttcgcctgag cggccgcgtc gacggaattc gcccttttcg
     4441 gatccactag taacggccgc cagtgtgctg gaattaattc gctgtctgcg agggccagct
     4501 gttggggtga gtactccctc tcaaaagcgg gcatgacttc tgcgctaaga ttgtcagttt
     4561 ccaaaaacga ggaggatttg atattcacct ggcccgcggt gatgcctttg agggtggccg
     4621 cgtccatctg gtcagaaaag acaatctttt tgttgtcaag cttgaggtgt ggcaggcttg
     4681 agatctggcc atacacttga gtgacaatga catccacttt gcctttctct ccacaggtgt
     4741 ccactcccag gtccaactgc aggtcgagca tgcatctagg gcggccaatt ccgcccctct
     4801 ccctcccccc cccctaacgt tactggccga agccgcttgg aataaggccg gtgtgcgttt
     4861 gtctatatgt gattttccac catattgccg tcttttggca atgtgagggc ccggaaacct
     4921 ggccctgtct tcttgacgag cattcctagg ggtctttccc ctctcgccaa aggaatgcaa
     4981 ggtctgttga atgtcgtgaa ggaagcagtt cctctggaag cttcttgaag acaaacaacg
     5041 tctgtagcga ccctttgcag gcagcggaac cccccacctg gcgacaggtg cctctgcggc
     5101 caaaagccac gtgtataaga tacacctgca aaggcggcac aaccccagtg ccacgttgtg
     5161 agttggatag ttgtggaaag agtcaaatgg ctctcctcaa gcgtattcaa caaggggctg
     5221 aaggatgccc agaaggtacc ccattgtatg ggatctgatc tggggcctcg gtgcacatgc
     5281 tttacatgtg tttagtcgag gttaaaaaaa cgtctaggcc ccccgaacca cggggacgtg
     5341 gttttccttt gaaaaacacg atgataagct tgccacaacc cacaaggaga cgaccttcca
     5401 tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc cgggccgtac
     5461 gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gacccggacc
     5521 gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca
     5581 tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga
     5641 gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt
     5701 cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc
     5761 ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca
     5821 gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg
     5881 agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg
     5941 acgtcgagtg cccgaaggac cgcgcgacct ggtgcatgac ccgcaagccc ggtgcctgac
     6001 gcccgcccca cgacccgcag cgcccgaccg aaaggagcgc acgaccccat ggctccgacc
     6061 gaagccgacc cgggcggccc cgccgacccc gcacccgccc ccgaggccca ccgactctag
     6121 agctcgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     6181 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     6241 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     6301 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     6361 tatggcttct gaggcggaaa gaaccagctg gggctcgagt gcattctagt tgtggtttgt
     6421 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg
     6481 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     6541 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     6601 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     6661 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     6721 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     6781 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     6841 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     6901 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     6961 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     7021 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     7081 tttctcaatg ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     7141 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     7201 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     7261 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     7321 gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     7381 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     7441 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     7501 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     7561 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     7621 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     7681 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     7741 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     7801 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     7861 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     7921 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     7981 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     8041 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     8101 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     8161 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     8221 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     8281 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     8341 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     8401 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     8461 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     8521 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     8581 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     8641 ctgacgtc
//
