LOCUS       sequence-363640-        7759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a Cmtr2-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-sgCmtr2#2/Cre
FEATURES             Location/Qualifiers
     source          1..7759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(17..36)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        208..587
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        589..787
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     742..762
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             805..985
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1032..1157
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1654..1887
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2072..2116
                     /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    2414..2531
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2613..2861
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2613..2633
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2784..2803
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2882..2957
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3010..3509
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3045..3068)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3442..3461
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3513..3533
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3534..4562
                     /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    4618..5206
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4671..4691)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5089..5100)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5209..5225)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5210..5226)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5735..5915
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5977..6565)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6057..6076)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6736..7527,7528..7596))
                     /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     7359..7378
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7597..7701)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatctgctcc
       61 ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct acaacaaggc
      121 aaggcttgac cgacaattgc atgaagaatc tgcttagggt taggcgtttt gcgctgcttc
      181 gcgatgtacg ggccagatat acgcgttgac attgattatt gactagttat taatagtaat
      241 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
      301 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
      361 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
      421 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
      481 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
      541 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
      601 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
      661 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
      721 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
      781 taagcagcgc gttttgcctg tactgggtct ctctggttag accagatctg agcctgggag
      841 ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc ttgagtgctt
      901 caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct cagacccttt
      961 tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa gcgaaaggga
     1021 aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg gcaagaggcg
     1081 aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag aaggagagag
     1141 atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg gaaaaaattc
     1201 ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg gcaagcaggg
     1261 agctagaacg attcgcagtt aatcctggcc tgttagaaac atcagaaggc tgtagacaaa
     1321 tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga tcattatata
     1381 atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac accaaggaag
     1441 ctttagacaa gatagaggaa gagcaaaaca aaagtaagac caccgcacag caagcggccg
     1501 gccgctgatc ttcagacctg gaggaggaga tatgagggac aattggagaa gtgaattata
     1561 taaatataaa gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg caaagagaag
     1621 agtggtgcag agagaaaaaa gagcagtggg aataggagct ttgttccttg ggttcttggg
     1681 agcagcagga agcactatgg gcgcagcgtc aatgacgctg acggtacagg ccagacaatt
     1741 attgtctggt atagtgcagc agcagaacaa tttgctgagg gctattgagg cgcaacagca
     1801 tctgttgcaa ctcacagtct ggggcatcaa gcagctccag gcaagaatcc tggctgtgga
     1861 aagataccta aaggatcaac agctcctggg gatttggggt tgctctggaa aactcatttg
     1921 caccactgct gtgccttgga atgctagttg gagtaataaa tctctggaac agatttggaa
     1981 tcacacgacc tggatggagt gggacagaga aattaacaat tacacaagct taatacactc
     2041 cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa caagaattat tggaattaga
     2101 taaatgggca agtttgtgga attggtttaa cataacaaat tggctgtggt atataaaatt
     2161 attcataatg atagtaggag gcttggtagg tttaagaata gtttttgctg tactttctat
     2221 agtgaataga gttaggcagg gatattcacc attatcgttt cagacccacc tcccaacccc
     2281 gaggggaccc gacaggcccg aaggaataga agaagaaggt ggagagagag acagagacag
     2341 atccattcga ttagtgaacg gatcggcact gcgtgcgcca attctgcaga caaatggcag
     2401 tattcatcca caattttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa
     2461 tagtagacat aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa
     2521 ttcaaaattt tcgggtttat tacagggaca gcagagatcc agtttggtta gtaccgggcc
     2581 cgctctagcc ctgcaggaac tgtttaaact gtgagggcct atttcccatg attccttcat
     2641 atttgcatat acgatacaag gctgttagag agataattgg aattaatttg actgtaaaca
     2701 caaagatatt agtacaaaat acgtgacgta gaaagtaata atttcttggg tagtttgcag
     2761 ttttaaaatt atgttttaaa atggactatc atatgcttac cgtaacttga aagtatttcg
     2821 atttcttggc tttatatatc ttgtggaaag gaccaaacac cgtaagccac tcgataatga
     2881 ggttttagag ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag
     2941 tggcaccgag tcggtgcttt tttgtttggc gcgcctatgg cttaattaac atggcggccg
     3001 cattctaccg ggtaggggag gcgcttttcc caaggcagtc tggagcatgc gctttagcag
     3061 ccccgctggg cacttggcgc tacacaagtg gcctctggcc tcgcacacat tccacatcca
     3121 ccggtaggcg ccaaccggct ccgttctttg gtggcccctt cgcgccacct tctactcctc
     3181 ccctagtcag gaagttcccc cccgccccgc agctcgcgtc gtgcaggacg tgacaaatgg
     3241 aagtagcacg tctcactagt ctcgtgcaga tggacagcac cgctgagcaa tggaagcggg
     3301 taggcctttg gggcagcggc caatagcagc tttgctcctt cgctttctgg gctcagaggc
     3361 tgggaagggg tgggtccggg ggcgggctca ggggcgggct caggggcggg gcgggcgccc
     3421 gaaggtcctc cggaggcccg gcattctgca cgcttcaaaa gcgcacgtct gccgcgctgt
     3481 tctcctcttc ctcatctccg ggacccggga tgcccaagaa gaagaggaag gtgtccaatt
     3541 tactgaccgt acaccaaaat ttgcctgcat taccggtcga tgcaacgagt gatgaggttc
     3601 gcaagaacct gatggacatg ttcagggatc gccaggcgtt ttctgagcat acctggaaaa
     3661 tgcttctgtc cgtttgccgg tcgtgggcgg catggtgcaa gttgaataac cggaaatggt
     3721 ttcccgcaga acctgaagat gttcgcgatt atcttctata tcttcaggcg cgcggtctgg
     3781 cagtaaaaac tatccagcaa catttgggcc agctaaacat gcttcatcgt cggtccgggc
     3841 tgccacgacc aagtgacagc aatgctgttt cactggttat gcggcggatc cgaaaagaaa
     3901 acgttgatgc cggtgaacgt gcaaaacagg ctctagcgtt cgaacgcact gatttcgacc
     3961 aggttcgttc actcatggaa aatagcgatc gctgccagga tatacgtaat ctggcatttc
     4021 tggggattgc ttataacacc ctgttacgta tagccgaaat tgccaggatc agggttaaag
     4081 atatctcacg tactgacggt gggagaatgt taatccatat tggcagaacg aaaacgctgg
     4141 ttagcaccgc aggtgtagag aaggcactta gcctgggggt aactaaactg gtcgagcgat
     4201 ggatttccgt ctctggtgta gctgatgatc cgaataacta cctgttttgc cgggtcagaa
     4261 aaaatggtgt tgccgcgcca tctgccacca gccagctatc aactcgcgcc ctggaaggga
     4321 tttttgaagc aactcatcga ttgatttacg gcgctaagga tgactctggt cagagatacc
     4381 tggcctggtc tggacacagt gcccgtgtcg gagccgcgcg agatatggcc cgcgctggag
     4441 tttcaatacc ggagatcatg caagctggtg gctggaccaa tgtaaatatt gtcatgaact
     4501 atatccgtaa cctggatagt gaaacagggg caatggtgcg cctgctggaa gatggcgatt
     4561 agccatgttt aagggttccg gttccactag gtacaattcg atatcaagct tatcgataat
     4621 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     4681 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     4741 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg
     4801 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt
     4861 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt
     4921 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     4981 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc
     5041 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat
     5101 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc
     5161 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgtcga
     5221 cctcgatcga gacctagaaa aacatggagc aatcacaagt agcaatacag cagctaccaa
     5281 tgctgattgt gcctggctag aagcacaaga ggaggaggag gtgggttttc cagtcacacc
     5341 tcaggtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa
     5401 agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagata tccttgatct
     5461 gtggatctac cacacacaag gctacttccc tgattggcag aactacacac cagggccagg
     5521 gatcagatat ccactgacct ttggatggtg ctacaagcta gtaccagttg agcaagagaa
     5581 ggtagaagaa gccaatgaag gagagaacac ccgcttgtta caccctgtga gcctgcatgg
     5641 gatggatgac ccggagagag aagtattaga gtggaggttt gacagccgcc tagcatttca
     5701 tcacatggcc cgagagctgc atccggactg tactgggtct ctctggttag accagatctg
     5761 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
     5821 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
     5881 cagacccttt tagtcagtgt ggaaaatctc tagcagcatg tgagcaaaag gccagcaaaa
     5941 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     6001 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     6061 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     6121 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     6181 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     6241 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     6301 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     6361 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     6421 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     6481 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6541 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6601 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6661 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6721 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6781 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6841 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6901 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6961 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     7021 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     7081 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     7141 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     7201 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     7261 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     7321 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     7381 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     7441 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7501 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7561 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7621 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7681 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
     7741 gggagatctc ccgatcccc
//
