LOCUS       sequence-218564-        7326 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of human PNGase with C-terminal strep-HA tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7326)
  AUTHORS   Hulsmann J, Kravic B, Weith M, Gstaiger M, Aebersold RH, Collins BC,
            Meyer H
  TITLE     AP-SWATH reveals direct involvement of VCP/p97 in integrated stress 
            response signaling through facilitating CReP/PPP1R15B degradation.
  JOURNAL   Mol Cell Proteomics. 2018 Mar 29. pii: RA117.000471. doi: 
            10.1074/mcp.RA117.000471.
  PUBMED    29599191
REFERENCE   2  (bases 1 to 7326)
  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: "Mol Cell 
            Proteomics. 2018 Mar 29. pii: RA117.000471. doi: 
            10.1074/mcp.RA117.000471."
COMMENT     Sequence Label:  pcDNA5FRT/TO-PNGase-Strep-HA
FEATURES             Location/Qualifiers
     source          1..7326
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        66..445
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        446..649
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     600..620
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    651..669
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    672..690
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     700..724
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    complement(2854..2878)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2925..2948
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             2985..3008
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             3039..3065
                     /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             3039..3065
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(3109..3126)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3115..3339
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3472..3491)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3623..3670
                     /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             3678..4700
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3678..4700
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4830..4951
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4867..4886)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4921..4940
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5000..5016)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5000..5016)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5013..5035)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5024..5040
                     /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(5048..5054,5055..5072,5073..5078))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5093..5114
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5231..5248)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5402..5990)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5482..5501)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6161..6952,6953..7021))
                     /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     6784..6803
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7022..7126)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7201..7220)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac
       61 gcgttgacat tgattattga ctagttatta atagtaatca attacggggt cattagttca
      121 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
      181 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
      241 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
      301 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
      361 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
      421 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
      481 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
      541 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
      601 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc tccctatcag
      661 tgatagagat ctccctatca gtgatagaga tcgtcgacga gctcgtttag tgaaccgtca
      721 gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagacacc gggaccgatc
      781 cagcctccgg actctagcgt ttaaacttaa gcttggtacc gagctcggat ccactagtcc
      841 agtgtggtgg aattctgcag atatcaacaa gtttgtacca aaaagttggc atggcggcgg
      901 cggcattggg cagctcctca ggctcggcgt ccccggccgt ggctgagctc tgccagaaca
      961 ccccggagac ctttttggag gcctccaagc tgctgctcac ctatgctgac aacatcctca
     1021 gaaaccctaa tgatgaaaaa tatagatcca tccggattgg aaacacagcc ttttctacta
     1081 gactcttgcc tgtcagagga gctgttgaat gtttatttga aatgggcttt gaagagggag
     1141 aaacacatct catctttcct aaaaaagctt cagtggagca gctgcaaaaa attcgtgacc
     1201 tgattgccat agagagaagt agcagactgg atggctcaaa taagagccac aaagtaaagt
     1261 catctcagca acctgcagcc agtacccagc ttcctacaac accatcttca aatcccagtg
     1321 ggttaaacca gcacacaagg aaccgtcaag ggcagtcatc agatccacca tctgcttcaa
     1381 cggttgctgc tgactcagcc attctagaag ttcttcagtc caacattcag catgtgctgg
     1441 tctatgaaaa tcctgctctt caggagaaag cgttggcttg tattccggtc caagaactaa
     1501 aaaggaaatc acaagaaaag ttatcgagag ctagaaaatt ggataaaggt atcaatataa
     1561 gtgatgagga ttttcttttg ctggagcttt tgcactggtt taaggaagaa ttttttcact
     1621 gggtgaataa cgttttgtgc agcaaatgtg gtggacagac taggtctaga gatagatcat
     1681 tactgcccag tgatgatgag ctgaagtggg gtgcaaagga agtggaagat cattactgtg
     1741 atgcctgcca gttcagcaat cgattcccaa gatataataa ccctgagaaa cttttggaaa
     1801 caagatgtgg acggtgtggc gagtgggcca attgttttac actgtgctgc cgagctgtag
     1861 ggtttgaagc tcgctatgtt tgggattaca cagaccatgt ctggacagaa gtctattctc
     1921 cttctcagca gcggtggctg cactgtgatg catgtgaaga tgtctgtgac aagccactcc
     1981 tttatgaaat aggatggggc aagaagcttt cctatgtcat agcattttca aaagatgagg
     2041 tagttgatgt cacttggcga tattcctgca aacatgaaga ggtgattgcc agaagaacta
     2101 aggttaaaga agcattactt cgagacacta ttaatgggct taataagcag aggcaactgt
     2161 ttttgtcaga aaacagaagg aaagaacttc tccagaggat aattgtggag cttgttgaat
     2221 ttatatctcc caaaacccct aaacctggag aacttggggg aagaatatct gggtcagtgg
     2281 cttggagagt agcccgaggt gaaatgggtc tacagagaaa agaaaccttg tttattccct
     2341 gtgaaaatga gaagatttct aaacagctcc acctttgtta caatattgtg aaagatcgtt
     2401 atgttcgagt ttcaaataac aatcaaacca tttctggatg ggagaatggc gtgtggaaaa
     2461 tggaatctat attcagaaaa gttgaaacag actggcacat ggtatatttg gcccgaaagg
     2521 aaggatcatc ttttgcttat atttcctgga agtttgagtg tgggtcagtt ggcctaaaag
     2581 tagatagcat ttctattaga acaagtagtc aaacttttca gactggaaca gtagaatgga
     2641 aattgcgatc tgatacagca caagtagaac tgacaggcga taacagtctt cactcctatg
     2701 ctgatttttc tggtgccact gaagttattt tggaagcaga attaagcaga ggagatggtg
     2761 atgtcgcttg gcaacacacc cagctgttta gacaaagctt aaatgaccat gaagaaaatt
     2821 gtttggagat aattataaaa ttcagtgacc tttgcccaac tttcttgtac aaagtggttg
     2881 atatcaaggc cgccgacatc accagcctgt acaagaaagt ggggtggagc catccccagt
     2941 tcgagaaggg cggaggatct ggcggcggaa gcggcggagg aagctggtcc caccctcagt
     3001 ttgagaaagg caccgagctg ggcagcacca tggccagcta cccctacgac gtgcccgact
     3061 acgcctgatg actcgagtct agagggcccg tttaaacccg ctgatcagcc tcgactgtgc
     3121 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
     3181 gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta
     3241 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
     3301 acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca
     3361 gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     3421 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     3481 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     3541 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     3601 agggtgatgg ttcacgtacc tagaagttcc tattccgaag ttcctattct ctagaaagta
     3661 taggaacttc cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag aagtttctga
     3721 tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa gaatctcgtg
     3781 ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc tgcgccgatg
     3841 gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc ccgattccgg
     3901 aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc cgccgtgcac
     3961 agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg cagccggtcg
     4021 cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg ttcggcccat
     4081 tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc gcgattgctg
     4141 atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg tccgtcgcgc
     4201 aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg cacctcgtgc
     4261 acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca gcggtcattg
     4321 actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc ttcttctgga
     4381 ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg catccggagc
     4441 ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac caactctatc
     4501 agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa
     4561 tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga agcgcggccg
     4621 tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc cccagcactc
     4681 gtccgagggc aaaggaatag cacgtactac gagatttcga ttccaccgcc gccttctatg
     4741 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     4801 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca
     4861 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     4921 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct
     4981 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     5041 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     5101 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     5161 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     5221 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     5281 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     5341 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     5401 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     5461 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     5521 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     5581 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     5641 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     5701 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     5761 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     5821 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     5881 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     5941 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     6001 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     6061 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     6121 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     6181 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     6241 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     6301 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     6361 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     6421 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     6481 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     6541 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6601 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6661 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6721 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     6781 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6841 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6901 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6961 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     7021 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     7081 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     7141 aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag
     7201 tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga
     7261 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa
     7321 ttgcat
//
