LOCUS       sequence-361624-        7759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a Atm-targeting gRNA and Cre-recombinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7759)
  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 7759)
  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-sgAtm#2/Cre
FEATURES             Location/Qualifiers
     source          1..7759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(104..123)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        295..674
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        676..874
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     829..849
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             892..1072
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1119..1244
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1741..1974
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2159..2203
                     /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    2501..2618
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2700..2948
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2700..2720
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2871..2890
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2969..3044
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3097..3596
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3132..3155)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3529..3548
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3600..3620
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3621..4649
                     /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    4705..5293
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4758..4778)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5176..5187)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5296..5312)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5297..5313)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5822..6002
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6064..6652)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6144..6163)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6823..7614,7615..7683))
                     /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     7446..7465
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7684..7759,1..29))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
       61 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
      121 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      181 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      241 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      301 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      361 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      421 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      481 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      541 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      601 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      661 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      721 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      781 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      841 gtaggcgtgt acggtgggag gtctatataa gcagcgcgtt ttgcctgtac tgggtctctc
      901 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
      961 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
     1021 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc
     1081 cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc
     1141 ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt
     1201 tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga
     1261 gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat
     1321 taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt
     1381 tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag
     1441 gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa
     1501 ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa
     1561 gtaagaccac cgcacagcaa gcggccggcc gctgatcttc agacctggag gaggagatat
     1621 gagggacaat tggagaagtg aattatataa atataaagta gtaaaaattg aaccattagg
     1681 agtagcaccc accaaggcaa agagaagagt ggtgcagaga gaaaaaagag cagtgggaat
     1741 aggagctttg ttccttgggt tcttgggagc agcaggaagc actatgggcg cagcgtcaat
     1801 gacgctgacg gtacaggcca gacaattatt gtctggtata gtgcagcagc agaacaattt
     1861 gctgagggct attgaggcgc aacagcatct gttgcaactc acagtctggg gcatcaagca
     1921 gctccaggca agaatcctgg ctgtggaaag atacctaaag gatcaacagc tcctggggat
     1981 ttggggttgc tctggaaaac tcatttgcac cactgctgtg ccttggaatg ctagttggag
     2041 taataaatct ctggaacaga tttggaatca cacgacctgg atggagtggg acagagaaat
     2101 taacaattac acaagcttaa tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa
     2161 gaatgaacaa gaattattgg aattagataa atgggcaagt ttgtggaatt ggtttaacat
     2221 aacaaattgg ctgtggtata taaaattatt cataatgata gtaggaggct tggtaggttt
     2281 aagaatagtt tttgctgtac tttctatagt gaatagagtt aggcagggat attcaccatt
     2341 atcgtttcag acccacctcc caaccccgag gggacccgac aggcccgaag gaatagaaga
     2401 agaaggtgga gagagagaca gagacagatc cattcgatta gtgaacggat cggcactgcg
     2461 tgcgccaatt ctgcagacaa atggcagtat tcatccacaa ttttaaaaga aaagggggga
     2521 ttggggggta cagtgcaggg gaaagaatag tagacataat agcaacagac atacaaacta
     2581 aagaattaca aaaacaaatt acaaaaattc aaaattttcg ggtttattac agggacagca
     2641 gagatccagt ttggttagta ccgggcccgc tctagccctg caggaactgt ttaaactgtg
     2701 agggcctatt tcccatgatt ccttcatatt tgcatatacg atacaaggct gttagagaga
     2761 taattggaat taatttgact gtaaacacaa agatattagt acaaaatacg tgacgtagaa
     2821 agtaataatt tcttgggtag tttgcagttt taaaattatg ttttaaaatg gactatcata
     2881 tgcttaccgt aacttgaaag tatttcgatt tcttggcttt atatatcttg tggaaaggac
     2941 caaacaccgt atctcagcaa cagtggctgt tttagagcta gaaatagcaa gttaaaataa
     3001 ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt gtttggcgcg
     3061 cctatggctt aattaacatg gcggccgcat tctaccgggt aggggaggcg cttttcccaa
     3121 ggcagtctgg agcatgcgct ttagcagccc cgctgggcac ttggcgctac acaagtggcc
     3181 tctggcctcg cacacattcc acatccaccg gtaggcgcca accggctccg ttctttggtg
     3241 gccccttcgc gccaccttct actcctcccc tagtcaggaa gttccccccc gccccgcagc
     3301 tcgcgtcgtg caggacgtga caaatggaag tagcacgtct cactagtctc gtgcagatgg
     3361 acagcaccgc tgagcaatgg aagcgggtag gcctttgggg cagcggccaa tagcagcttt
     3421 gctccttcgc tttctgggct cagaggctgg gaaggggtgg gtccgggggc gggctcaggg
     3481 gcgggctcag gggcggggcg ggcgcccgaa ggtcctccgg aggcccggca ttctgcacgc
     3541 ttcaaaagcg cacgtctgcc gcgctgttct cctcttcctc atctccggga cccgggatgc
     3601 ccaagaagaa gaggaaggtg tccaatttac tgaccgtaca ccaaaatttg cctgcattac
     3661 cggtcgatgc aacgagtgat gaggttcgca agaacctgat ggacatgttc agggatcgcc
     3721 aggcgttttc tgagcatacc tggaaaatgc ttctgtccgt ttgccggtcg tgggcggcat
     3781 ggtgcaagtt gaataaccgg aaatggtttc ccgcagaacc tgaagatgtt cgcgattatc
     3841 ttctatatct tcaggcgcgc ggtctggcag taaaaactat ccagcaacat ttgggccagc
     3901 taaacatgct tcatcgtcgg tccgggctgc cacgaccaag tgacagcaat gctgtttcac
     3961 tggttatgcg gcggatccga aaagaaaacg ttgatgccgg tgaacgtgca aaacaggctc
     4021 tagcgttcga acgcactgat ttcgaccagg ttcgttcact catggaaaat agcgatcgct
     4081 gccaggatat acgtaatctg gcatttctgg ggattgctta taacaccctg ttacgtatag
     4141 ccgaaattgc caggatcagg gttaaagata tctcacgtac tgacggtggg agaatgttaa
     4201 tccatattgg cagaacgaaa acgctggtta gcaccgcagg tgtagagaag gcacttagcc
     4261 tgggggtaac taaactggtc gagcgatgga tttccgtctc tggtgtagct gatgatccga
     4321 ataactacct gttttgccgg gtcagaaaaa atggtgttgc cgcgccatct gccaccagcc
     4381 agctatcaac tcgcgccctg gaagggattt ttgaagcaac tcatcgattg atttacggcg
     4441 ctaaggatga ctctggtcag agatacctgg cctggtctgg acacagtgcc cgtgtcggag
     4501 ccgcgcgaga tatggcccgc gctggagttt caataccgga gatcatgcaa gctggtggct
     4561 ggaccaatgt aaatattgtc atgaactata tccgtaacct ggatagtgaa acaggggcaa
     4621 tggtgcgcct gctggaagat ggcgattagc catgtttaag ggttccggtt ccactaggta
     4681 caattcgata tcaagcttat cgataatcaa cctctggatt acaaaatttg tgaaagattg
     4741 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     4801 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     4861 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     4921 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     4981 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     5041 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     5101 tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     5161 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     5221 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     5281 gccgcctccc cgcatcgata ccgtcgacct cgatcgagac ctagaaaaac atggagcaat
     5341 cacaagtagc aatacagcag ctaccaatgc tgattgtgcc tggctagaag cacaagagga
     5401 ggaggaggtg ggttttccag tcacacctca ggtaccttta agaccaatga cttacaaggc
     5461 agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc
     5521 ccaacgaaga caagatatcc ttgatctgtg gatctaccac acacaaggct acttccctga
     5581 ttggcagaac tacacaccag ggccagggat cagatatcca ctgacctttg gatggtgcta
     5641 caagctagta ccagttgagc aagagaaggt agaagaagcc aatgaaggag agaacacccg
     5701 cttgttacac cctgtgagcc tgcatgggat ggatgacccg gagagagaag tattagagtg
     5761 gaggtttgac agccgcctag catttcatca catggcccga gagctgcatc cggactgtac
     5821 tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc
     5881 actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt
     5941 gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag
     6001 cagcatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     6061 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     6121 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     6181 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     6241 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     6301 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     6361 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     6421 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     6481 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     6541 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     6601 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     6661 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     6721 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     6781 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     6841 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     6901 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     6961 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     7021 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     7081 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     7141 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     7201 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     7261 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     7321 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     7381 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     7441 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     7501 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     7561 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     7621 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     7681 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     7741 atacatattt gaatgtatt
//
