LOCUS       sequence-163082-        7031 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  rAAV-based template for genome engineering of the p53 C-terminus 
            containing PQS1 and 3xFLAG tags and a selection cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7031)
  AUTHORS   Vandemoortele G, Staes A, Gonnelli G, Samyn N, De Sutter D, 
            Vandermarliere E, Timmerman E, Gevaert K, Martens L, Eyckerman S
  TITLE     An extra dimension in protein tagging by quantifying universal 
            proteotypic peptides using targeted proteomics.
  JOURNAL   Sci Rep. 2016 Jun 6;6:27220. doi: 10.1038/srep27220.
  PUBMED    27264994
REFERENCE   2  (bases 1 to 7031)
  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: "Sci Rep."; 
            date: "2016-06-6"; volume: "6:27220. doi"; pages: " 
            10.1038/srep27220"
COMMENT     Sequence Label: pAav-TP53-PQS1-3xFLAG
FEATURES             Location/Qualifiers
     source          1..7031
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     protein_bind    1270..1303
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     CDS             1374..1439
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    complement(1467..1500)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1512..2012
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1547..1570)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1945..1964
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             2026..2829
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVNQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2089..2108)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2699..2718
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2861..2878)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2867..3091
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(3116..3149)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      4073..4082
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     repeat_region   4294..4434
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4294..4423
                     /label=AAV2 ITR
     rep_origin      4509..4964
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4596..4615)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4806..4827
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4981..5000)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5100..5122
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5160..5178)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5246..5350
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5351..5419,5420..6211)
                     /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     complement(5569..5588)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6382..6970
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6871..6890
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca aacgcgtgtt ccgagagctg aatgaggcct
      181 tggaactcaa ggatgcccag gctgggaagg agccaggggg gagcagggct cactccaggt
      241 gagtgacctc agccccttcc tggccctact cccctgcctt cctaggttgg aaagccatag
      301 gattccattc tcatcctgcc ttcatggtca aaggcagctg accccatctc attgggtccc
      361 agccctgcac agacattttt ttagtcttcc tccggttgaa tcctataacc acattcttgc
      421 ctcagtgtat ccacagaaca tccaaaccca gggacgagtg tggatacttc tttgccattc
      481 tccgcaactc ccagcccaga gctggagggt ctcaaggagg ggcctaataa ttgtgtaata
      541 ctgaatacag ccagagtttc aggtcatata ctcagccctg ccatgcaccg gcaggtccta
      601 ggtgaccccc gtcaaactca gtttccttat atataaaatg gggtaagggg gccgggcgca
      661 gtggctcacg aatcccacac tctgggaggc caaggcgagt ggatcacctg aggtcgggag
      721 tttgagccca gcctgaccaa catggagaaa ccccatctct actaaaaata caaaagtagc
      781 cgggcgtggt gatgcatgcc tgtaatccca gctacctact cgggaggctg aggcaggaga
      841 atcgcttgaa cccgggaggc agaggttgcg gtgagctgag atctcaccat tacactccag
      901 cctgggcaac aagagtgaaa ctccgtctca aaaaagataa ataaagtaaa atggggtaag
      961 ggaagattac gagactaata cacactaata ctctgaggtg ctcagtaaac atatttgcat
     1021 ggggtgtggc caccatcttg atttgaattc ccgttgtccc agccttaggc ccttcaaagc
     1081 attggtcagg gaaaaggggc acagaccctc tcactcatgt gatgtcatct ctcctccctg
     1141 cttctgtctc ctacagccac ctgaagtcca aaaagggtca gtctacctcc cgccataaaa
     1201 aactcatgtt caagacagaa gggcctgact cagacagcgg gcccagcgaa ttcgggaagt
     1261 tcctattccg aagttcctat tcttcaaata gtataggaac ttcactgacc ctgatcttca
     1321 gagaggccgt gagcgaaatc ctggagacct ccagattcgc ctacctgtac gacgactaca
     1381 aagaccatga cggtgattat aaagatcatg atattgatta caaggatgac gatgacaagg
     1441 gcgagcccga ggcttgaaag cttcggataa cttcgtatag catacattat acgaagttat
     1501 cgcgccctac cgggtagggg aggcgctttt cccaaggcag tctggagcat gcgctttagc
     1561 agccccgctg ggcacttggc gctacacaag tggcctctgg cctcgcacac attccacatc
     1621 caccggtagg cgccaaccgg ctccgttctt tggtggcccc ttcgcgccac cttctactcc
     1681 tcccctagtc aggaagttcc cccccgcccc gcagctcgcg tcgtgcagga cgtgacaaat
     1741 ggaagtagca cgtctcacta gtctcgtgca gatggacagc accgctgagc aatggaagcg
     1801 ggtaggcctt tggggcagcg gccaatagca gctttggctc cttcgctttc tgggctcaga
     1861 ggctgggaag gggtgggtcc gggggcgggc tcaggggcgg gctcaggggc ggggcgggcg
     1921 cccgaaggtc ctccggaagc ccggcattct gcacgcttca aaagcgcacg tctgccgcgc
     1981 tgttctcctc ttcctcatct ccgggccttt cgacctgcag ccaatatggg atcggccatt
     2041 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
     2101 gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag
     2161 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac
     2221 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
     2281 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
     2341 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
     2401 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
     2461 cgagcacgta ctcggatgga agccggtctt gtcaatcagg atgatctgga cgaagagcat
     2521 caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag
     2581 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
     2641 ttttctggat tcatcgactg tggccggctg ggtgtggcgg atcgctatca ggacatagcg
     2701 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
     2761 ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
     2821 ttcttctgag gggatcaatt ctctagagct cgctgatcag cctcgactgt gccttctagt
     2881 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     2941 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     3001 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     3061 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctataac
     3121 ttcgtatagc atacattata cgaagttata gtaatcgatt gacattctcc acttcttgtt
     3181 ccccactgac agcctcccac ccccatctct ccctcccctg ccattttggg ttttgggtct
     3241 ttgaaccctt gcttgcaata ggtgtgcgtc agaagcaccc aggacttcca tttgctttgt
     3301 cccggggctc cactgaacaa gttggcctgc actggtgttt tgttgtgggg aggaggatgg
     3361 ggagtaggac ataccagctt agattttaag gtttttactg tgagggatgt ttgggagatg
     3421 taagaaatgt tcttgcagtt aagggttagt ttacaatcag ccacattcta ggtaggggcc
     3481 cacttcaccg tactaaccag ggaagctgtc cctcactgtt gaattttctc taacttcaag
     3541 gcccatatct gtgaaatgct ggcatttgca cctacctcac agagtgcatt gtgagggtta
     3601 atgaaataat gtacatctgg ccttgaaacc accttttatt acatggggtc tagaacttga
     3661 cccccttgag ggtgcttgtt ccctctccct gttggtcggt gggttggtag tttctacagt
     3721 tgggcagctg gttaggtaga gggagttgtc aagtctctgc tggcccagcc aaaccctgtc
     3781 tgacaacctc ttggtgaacc ttagtaccta aaaggaaatc tcaccccatc ccacaccctg
     3841 gaggatttca tctcttgtat atgatgatct ggatccacca agacttgttt tatgctcagg
     3901 gtcaatttct tttttctttt tttttttttt ttttcttttt ctttgagact gggtctcgct
     3961 ttgttgccca ggctggagtg gagtggcgtg atcttggctt actgcagcct ttgcctcccc
     4021 ggctcgagca gtcctgcctc agcctccgga gtagctggga ccacaggttc atgccaccat
     4081 ggccagccaa cttttgcatg ttttgtagag atggggtctc acagtgttgc ccaggctggt
     4141 ctcaaactcc tgggctcagg cgatccacct gtctcagcct cccagagtgc tgggattaca
     4201 attgtgagcc accacgtcca gctggaaggg tcaacatctt ttacattctg caagcacatc
     4261 tgcatttggt accaagctag caaaagcggc cgcaggaacc cctagtgatg gagttggcca
     4321 ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc
     4381 cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc
     4441 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca
     4501 accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     4561 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     4621 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     4681 ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg
     4741 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     4801 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg gctattcttt
     4861 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     4921 aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag
     4981 tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga
     5041 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     5101 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg
     5161 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc
     5221 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     5281 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     5341 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     5401 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     5461 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     5521 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     5581 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     5641 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     5701 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     5761 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     5821 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     5881 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     5941 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     6001 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     6061 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     6121 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     6181 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     6241 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     6301 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     6361 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     6421 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     6481 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta
     6541 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     6601 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     6661 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     6721 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     6781 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     6841 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     6901 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     6961 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     7021 tgctcacatg t
//
