LOCUS       sequence-363591-        7760 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a Ep300-targeting gRNA and Cre-recombinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7760)
  AUTHORS   Cai H, Chew SK, Li C, Tsai MK, Andrejka L, Murray CW, Hughes NW, 
            Shuldiner EG, Ashkin EL, Tang R, Hung KL, Chen LC, Lee SYC, Yousefi 
            M, Lin WY, Kunder CA, Cong L, McFarland CD, Petrov DA, Swanton C, 
            Winslow MM
  TITLE     A functional taxonomy of tumor suppression in oncogenic KRAS-driven 
            lung cancer.
  JOURNAL   Cancer Discov. 2021 Feb 19. pii: 2159-8290.CD-20-1325. doi: 
            10.1158/2159-8290.CD-20-1325.
  PUBMED    33608386
REFERENCE   2  (bases 1 to 7760)
  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: "Cancer 
            Discov. 2021 Feb 19. pii: 2159-8290.CD-20-1325. doi: 
            10.1158/2159-8290.CD-20-1325."
COMMENT     Sequence Label: Lenti-sgEp300#1/Cre
FEATURES             Location/Qualifiers
     source          1..7760
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(135..154)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        326..705
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        707..905
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     860..880
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             923..1103
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1150..1275
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1772..2005
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2190..2234
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2532..2649
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2731..2979
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2731..2751
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2902..2921
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        3001..3076
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3129..3628
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3164..3187)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3561..3580
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3632..3652
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3653..4681
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="SNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKMLL
                     SVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGLP
                     RPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAFL
                     GIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVER
                     WISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQR
                     YLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLED
                     GD"
     misc_feature    4737..5325
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4790..4810)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5208..5219)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5328..5344)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5329..5345)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5854..6034
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6096..6684)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6176..6195)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6855..7646,7647..7715))
                     /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     7478..7497
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7716..7760,1..60))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
       61 cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta
      121 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
      181 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa
      241 ggcttgaccg acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc
      301 gatgtacggg ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca
      361 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
      421 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
      481 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
      541 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
      601 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
      661 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
      721 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
      781 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
      841 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
      901 agcagcgcgt tttgcctgta ctgggtctct ctggttagac cagatctgag cctgggagct
      961 ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca
     1021 agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta
     1081 gtcagtgtgg aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa
     1141 ccagaggagc tctctcgacg caggactcgg cttgctgaag cgcgcacggc aagaggcgag
     1201 gggcggcgac tggtgagtac gccaaaaatt ttgactagcg gaggctagaa ggagagagat
     1261 gggtgcgaga gcgtcagtat taagcggggg agaattagat cgcgatggga aaaaattcgg
     1321 ttaaggccag ggggaaagaa aaaatataaa ttaaaacata tagtatgggc aagcagggag
     1381 ctagaacgat tcgcagttaa tcctggcctg ttagaaacat cagaaggctg tagacaaata
     1441 ctgggacagc tacaaccatc ccttcagaca ggatcagaag aacttagatc attatataat
     1501 acagtagcaa ccctctattg tgtgcatcaa aggatagaga taaaagacac caaggaagct
     1561 ttagacaaga tagaggaaga gcaaaacaaa agtaagacca ccgcacagca agcggccggc
     1621 cgctgatctt cagacctgga ggaggagata tgagggacaa ttggagaagt gaattatata
     1681 aatataaagt agtaaaaatt gaaccattag gagtagcacc caccaaggca aagagaagag
     1741 tggtgcagag agaaaaaaga gcagtgggaa taggagcttt gttccttggg ttcttgggag
     1801 cagcaggaag cactatgggc gcagcgtcaa tgacgctgac ggtacaggcc agacaattat
     1861 tgtctggtat agtgcagcag cagaacaatt tgctgagggc tattgaggcg caacagcatc
     1921 tgttgcaact cacagtctgg ggcatcaagc agctccaggc aagaatcctg gctgtggaaa
     1981 gatacctaaa ggatcaacag ctcctgggga tttggggttg ctctggaaaa ctcatttgca
     2041 ccactgctgt gccttggaat gctagttgga gtaataaatc tctggaacag atttggaatc
     2101 acacgacctg gatggagtgg gacagagaaa ttaacaatta cacaagctta atacactcct
     2161 taattgaaga atcgcaaaac cagcaagaaa agaatgaaca agaattattg gaattagata
     2221 aatgggcaag tttgtggaat tggtttaaca taacaaattg gctgtggtat ataaaattat
     2281 tcataatgat agtaggaggc ttggtaggtt taagaatagt ttttgctgta ctttctatag
     2341 tgaatagagt taggcaggga tattcaccat tatcgtttca gacccacctc ccaaccccga
     2401 ggggacccga caggcccgaa ggaatagaag aagaaggtgg agagagagac agagacagat
     2461 ccattcgatt agtgaacgga tcggcactgc gtgcgccaat tctgcagaca aatggcagta
     2521 ttcatccaca attttaaaag aaaagggggg attggggggt acagtgcagg ggaaagaata
     2581 gtagacataa tagcaacaga catacaaact aaagaattac aaaaacaaat tacaaaaatt
     2641 caaaattttc gggtttatta cagggacagc agagatccag tttggttagt accgggcccg
     2701 ctctagccct gcaggaactg tttaaactgt gagggcctat ttcccatgat tccttcatat
     2761 ttgcatatac gatacaaggc tgttagagag ataattggaa ttaatttgac tgtaaacaca
     2821 aagatattag tacaaaatac gtgacgtaga aagtaataat ttcttgggta gtttgcagtt
     2881 ttaaaattat gttttaaaat ggactatcat atgcttaccg taacttgaaa gtatttcgat
     2941 ttcttggctt tatatatctt gtggaaagga ccaaacaccg aagctcgctc tatagagctg
     3001 gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt
     3061 ggcaccgagt cggtgctttt ttgtttggcg cgcctatggc ttaattaaca tggcggccgc
     3121 attctaccgg gtaggggagg cgcttttccc aaggcagtct ggagcatgcg ctttagcagc
     3181 cccgctgggc acttggcgct acacaagtgg cctctggcct cgcacacatt ccacatccac
     3241 cggtaggcgc caaccggctc cgttctttgg tggccccttc gcgccacctt ctactcctcc
     3301 cctagtcagg aagttccccc ccgccccgca gctcgcgtcg tgcaggacgt gacaaatgga
     3361 agtagcacgt ctcactagtc tcgtgcagat ggacagcacc gctgagcaat ggaagcgggt
     3421 aggcctttgg ggcagcggcc aatagcagct ttgctccttc gctttctggg ctcagaggct
     3481 gggaaggggt gggtccgggg gcgggctcag gggcgggctc aggggcgggg cgggcgcccg
     3541 aaggtcctcc ggaggcccgg cattctgcac gcttcaaaag cgcacgtctg ccgcgctgtt
     3601 ctcctcttcc tcatctccgg gacccgggat gcccaagaag aagaggaagg tgtccaattt
     3661 actgaccgta caccaaaatt tgcctgcatt accggtcgat gcaacgagtg atgaggttcg
     3721 caagaacctg atggacatgt tcagggatcg ccaggcgttt tctgagcata cctggaaaat
     3781 gcttctgtcc gtttgccggt cgtgggcggc atggtgcaag ttgaataacc ggaaatggtt
     3841 tcccgcagaa cctgaagatg ttcgcgatta tcttctatat cttcaggcgc gcggtctggc
     3901 agtaaaaact atccagcaac atttgggcca gctaaacatg cttcatcgtc ggtccgggct
     3961 gccacgacca agtgacagca atgctgtttc actggttatg cggcggatcc gaaaagaaaa
     4021 cgttgatgcc ggtgaacgtg caaaacaggc tctagcgttc gaacgcactg atttcgacca
     4081 ggttcgttca ctcatggaaa atagcgatcg ctgccaggat atacgtaatc tggcatttct
     4141 ggggattgct tataacaccc tgttacgtat agccgaaatt gccaggatca gggttaaaga
     4201 tatctcacgt actgacggtg ggagaatgtt aatccatatt ggcagaacga aaacgctggt
     4261 tagcaccgca ggtgtagaga aggcacttag cctgggggta actaaactgg tcgagcgatg
     4321 gatttccgtc tctggtgtag ctgatgatcc gaataactac ctgttttgcc gggtcagaaa
     4381 aaatggtgtt gccgcgccat ctgccaccag ccagctatca actcgcgccc tggaagggat
     4441 ttttgaagca actcatcgat tgatttacgg cgctaaggat gactctggtc agagatacct
     4501 ggcctggtct ggacacagtg cccgtgtcgg agccgcgcga gatatggccc gcgctggagt
     4561 ttcaataccg gagatcatgc aagctggtgg ctggaccaat gtaaatattg tcatgaacta
     4621 tatccgtaac ctggatagtg aaacaggggc aatggtgcgc ctgctggaag atggcgatta
     4681 gccatgttta agggttccgg ttccactagg tacaattcga tatcaagctt atcgataatc
     4741 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
     4801 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
     4861 ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc
     4921 ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt
     4981 ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg
     5041 ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg
     5101 gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct
     5161 gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc
     5221 cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc
     5281 ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcatcga taccgtcgac
     5341 ctcgatcgag acctagaaaa acatggagca atcacaagta gcaatacagc agctaccaat
     5401 gctgattgtg cctggctaga agcacaagag gaggaggagg tgggttttcc agtcacacct
     5461 caggtacctt taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa
     5521 gaaaaggggg gactggaagg gctaattcac tcccaacgaa gacaagatat ccttgatctg
     5581 tggatctacc acacacaagg ctacttccct gattggcaga actacacacc agggccaggg
     5641 atcagatatc cactgacctt tggatggtgc tacaagctag taccagttga gcaagagaag
     5701 gtagaagaag ccaatgaagg agagaacacc cgcttgttac accctgtgag cctgcatggg
     5761 atggatgacc cggagagaga agtattagag tggaggtttg acagccgcct agcatttcat
     5821 cacatggccc gagagctgca tccggactgt actgggtctc tctggttaga ccagatctga
     5881 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
     5941 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
     6001 agaccctttt agtcagtgtg gaaaatctct agcagcatgt gagcaaaagg ccagcaaaag
     6061 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     6121 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     6181 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     6241 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     6301 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6361 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6421 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6481 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     6541 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6601 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6661 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     6721 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     6781 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     6841 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     6901 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     6961 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     7021 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     7081 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     7141 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     7201 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     7261 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     7321 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     7381 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     7441 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     7501 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     7561 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     7621 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     7681 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     7741 agcatttatc agggttattg
//
