LOCUS       sequence-379336-        5168 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Cre cDNA and sgRNAs targeting murine Trp53 and Atrx.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5168)
  AUTHORS   Shi DD, Savani MR, Levitt MM, Wang AC, Endress JE, Bird CE, Buehler 
            J, Stopka SA, Regan MS, Lin YF, Puliyappadamba VT, Gao W, Khanal J, 
            Evans L, Lee JH, Guo L, Xiao Y, Xu M, Huang B, Jennings RB, Bonal 
            DM, Martin-Sandoval MS, Dang T, Gattie LC, Cameron AB, Lee S, Asara 
            JM, Kornblum HI, Mak TW, Looper RE, Nguyen QD, Signoretti S, Gradl 
            S, Sutter A, Jeffers M, Janzer A, Lehrman MA, Zacharias LG, Mathews 
            TP, Losman JA, Richardson TE, Cahill DP, DeBerardinis RJ, Ligon KL, 
            Xu L, Ly P, Agar NYR, Abdullah KG, Harris IS, Kaelin WG Jr, McBrayer
            SK
  TITLE     De novo pyrimidine synthesis is a targetable vulnerability in IDH 
            mutant glioma.
  JOURNAL   Cancer Cell. 2022 Sep 12;40(9):939-956.e16. doi: 
            10.1016/j.ccell.2022.07.011. Epub 2022 Aug 18.
  PUBMED    35985343
REFERENCE   2  (bases 1 to 5168)
  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: 
            "10.1016/j.ccell.2022.07.011"; journalName: "Cancer Cell"; date: 
            "2022-09-12- 12"; volume: "40"; issue: "9"; pages: "939-956.e16"
COMMENT     Sequence Label: pAAV2-sgTrp53-sgAtrx-EFS-Cre
FEATURES             Location/Qualifiers
     source          1..5168
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(81..105)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     repeat_region   142..282
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   142..271
                     /label=AAV2 ITR
     rep_origin      357..812
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(444..463)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     654..675
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(829..848)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     948..970
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1008..1026)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1094..1198
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1199..1267,1268..2059)
                     /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(1417..1436)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2230..2818
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2719..2738
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   2880..3009
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     CDS             complement(3079..3105)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(4132..4152)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        complement(4157..4368)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     protein_bind    4440..4464
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_RNA        4726..4801
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    4848..4868
                     /gene="mutant version of attB"
                     /label=attB5
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     misc_RNA        join(5128..5168,1..35)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
ORIGIN
        1 gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt tgaattctag atcttgagac
       61 aaatggcagt attcatccac acccagcttt cttgtacaaa gtggtgggtg gaggcggttc
      121 aggtggcgga ggaagggccg caggaacccc tagtgatgga gttggccact ccctctctgc
      181 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
      241 gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt
      301 ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc
      361 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
      421 acttgccagc gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
      481 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
      541 tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc
      601 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
      661 cttgttccaa actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg
      721 gattttgccg atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc
      781 gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc
      841 tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg
      901 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
      961 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg
     1021 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     1081 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     1141 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     1201 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     1261 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     1321 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     1381 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     1441 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     1501 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     1561 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     1621 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     1681 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     1741 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     1801 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     1861 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtggaagccg
     1921 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     1981 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     2041 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     2101 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     2161 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     2221 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     2281 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     2341 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     2401 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     2461 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     2521 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     2581 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     2641 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     2701 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     2761 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     2821 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtc
     2881 ctgcaggcag ctgcgcgctc gctcgctcac tgaggccgcc cgggcgtcgg gcgacctttg
     2941 gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg agtggccaac tccatcacta
     3001 ggggttcctg cggcctctag aacacaaaaa accaacacac agatctaatg aaaataaaga
     3061 tcttttatta gcgctttaag cgtaatctgg aacatcgtat gggtagtcac catcttcgag
     3121 cagtctcacc atggccccgg tttcagagtc aagattgcgg atatagttca taacgatgtt
     3181 cacatttgtc caacctccag cttgcataat ctctggtatg gagactcctg cccgcgccat
     3241 gtctcgtgcg gcccccaccc tagcagaatg acctgaccag gcgagatacc tctggccact
     3301 gtcatccttg gctccgtata tgagtcggtg ggttgcctca aagatccctt caagggcccg
     3361 tgtactcaat tggctggtgg cagagggcgc ggcgactccg ttttttctca ctcgacagaa
     3421 caagtaatta ttagggtcgt cagccacgcc ggacactgag atccatcttt ccaccagctt
     3481 cgttacgccc agggacaagg ccttttcaac gccagcggtg ctgaccaatg tctttgtgcg
     3541 tccaatatga ataagcatcc ggccgccgtc cgttcgactg atgtctttca ctctgatccg
     3601 ggcgatctca gctatgcgca gaagtgtgtt atatgcaatt cccagaaatg cgaggttcct
     3661 gatgtcctgg caccgatcgc tgttctccat cagggatctg acctggtcga aatcagtccg
     3721 ctcaaaagcg agagcctgct tagccctctc acccgcatct acgttttcct ttctaattct
     3781 cctcataaca agggagactg cgtttgaatc gctaggccgg gggagcccgc tgcgtcgatg
     3841 cagcatattc agttggccca agtgctgctg gatagtcttg acagccagcc ctcttgcctg
     3901 gagatacagc aggtaatcgc gcacgtcctc cggctcggcg gggaaccatt tgcgattgtt
     3961 cagcttacac caggctgccc aggatcggca cacgctaagc agcattttcc aggtgtgttc
     4021 agaaaacgcc tgtctatccc taaacatatc catgagattt ttcctgacct catcgctagt
     4081 ggcatctact ggcagcgcag ggaggttctg gtgaacagtc aggagattgg acaccttcct
     4141 cttcttcttg ggcatcctgt gttctggcgg caaacccgtt gcgaaaaaga acgttcacgg
     4201 cgactactgc acttatatac ggttctcccc caccctcggg aaaaaggcgg agccagtaca
     4261 cgacatcact ttcccagttt accccgcgcc accttctcta ggcaccggtt caattgccga
     4321 cccctccccc caacttctcg gggactgtgg gcgatgtgcg ctctgcccac tgacgggcac
     4381 cggagccaat tcccactcct ttcaagacct ttaattaacc cacaaggaga cgaccttcca
     4441 caagtttgta caaaaaagca ggcttttccc atgattcctt catatttgca tatacgatac
     4501 aaggctgtta gagagataat tagaattaat ttgactgtaa acacaaagat attagtacaa
     4561 aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa attatgtttt
     4621 aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt ggctttatat
     4681 atcttgtgga aaggacgaaa caccggtgta atagctcctg catgggtttt agagctagaa
     4741 atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg
     4801 cttttttgaa ttctagatct tgagacaaat ggcagtattc atccacacaa ctttgtatac
     4861 aaaagttgtt tcccatgatt ccttcatatt tgcatatacg atacaaggct gttagagaga
     4921 taattagaat taatttgact gtaaacacaa agatattagt acaaaatacg tgacgtagaa
     4981 agtaataatt tcttgggtag tttgcagttt taaaattatg ttttaaaatg gactatcata
     5041 tgcttaccgt aacttgaaag tatttcgatt tcttggcttt atatatcttg tggaaaggac
     5101 gaaacaccga taaggacttt gagtgaagtt ttagagctag aaatagcaag ttaaaataag
     5161 gctagtcc
//
