LOCUS       sequence-363621-        7760 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a Setd2-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-sgSetd2#2/Cre
FEATURES             Location/Qualifiers
     source          1..7760
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(81..100)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        272..651
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        653..851
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     806..826
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             869..1049
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1096..1221
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1718..1951
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2136..2180
                     /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    2478..2595
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2677..2925
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2677..2697
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2848..2867
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2947..3022
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3075..3574
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3110..3133)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3507..3526
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3578..3598
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3599..4627
                     /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    4683..5271
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4736..4756)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5154..5165)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5274..5290)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5275..5291)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5800..5980
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6042..6630)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6122..6141)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6801..7592,7593..7661))
                     /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     7424..7443
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7662..7760,1..6))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat
       61 cccctatggt gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg
      121 ctccctgctt gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca
      181 aggcaaggct tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg
      241 cttcgcgatg tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag
      301 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
      361 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
      421 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
      481 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
      541 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
      601 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
      661 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
      721 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
      781 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
      841 tatataagca gcgcgttttg cctgtactgg gtctctctgg ttagaccaga tctgagcctg
      901 ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt
      961 gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc
     1021 cttttagtca gtgtggaaaa tctctagcag tggcgcccga acagggactt gaaagcgaaa
     1081 gggaaaccag aggagctctc tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga
     1141 ggcgaggggc ggcgactggt gagtacgcca aaaattttga ctagcggagg ctagaaggag
     1201 agagatgggt gcgagagcgt cagtattaag cgggggagaa ttagatcgcg atgggaaaaa
     1261 attcggttaa ggccaggggg aaagaaaaaa tataaattaa aacatatagt atgggcaagc
     1321 agggagctag aacgattcgc agttaatcct ggcctgttag aaacatcaga aggctgtaga
     1381 caaatactgg gacagctaca accatccctt cagacaggat cagaagaact tagatcatta
     1441 tataatacag tagcaaccct ctattgtgtg catcaaagga tagagataaa agacaccaag
     1501 gaagctttag acaagataga ggaagagcaa aacaaaagta agaccaccgc acagcaagcg
     1561 gccggccgct gatcttcaga cctggaggag gagatatgag ggacaattgg agaagtgaat
     1621 tatataaata taaagtagta aaaattgaac cattaggagt agcacccacc aaggcaaaga
     1681 gaagagtggt gcagagagaa aaaagagcag tgggaatagg agctttgttc cttgggttct
     1741 tgggagcagc aggaagcact atgggcgcag cgtcaatgac gctgacggta caggccagac
     1801 aattattgtc tggtatagtg cagcagcaga acaatttgct gagggctatt gaggcgcaac
     1861 agcatctgtt gcaactcaca gtctggggca tcaagcagct ccaggcaaga atcctggctg
     1921 tggaaagata cctaaaggat caacagctcc tggggatttg gggttgctct ggaaaactca
     1981 tttgcaccac tgctgtgcct tggaatgcta gttggagtaa taaatctctg gaacagattt
     2041 ggaatcacac gacctggatg gagtgggaca gagaaattaa caattacaca agcttaatac
     2101 actccttaat tgaagaatcg caaaaccagc aagaaaagaa tgaacaagaa ttattggaat
     2161 tagataaatg ggcaagtttg tggaattggt ttaacataac aaattggctg tggtatataa
     2221 aattattcat aatgatagta ggaggcttgg taggtttaag aatagttttt gctgtacttt
     2281 ctatagtgaa tagagttagg cagggatatt caccattatc gtttcagacc cacctcccaa
     2341 ccccgagggg acccgacagg cccgaaggaa tagaagaaga aggtggagag agagacagag
     2401 acagatccat tcgattagtg aacggatcgg cactgcgtgc gccaattctg cagacaaatg
     2461 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa
     2521 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca
     2581 aaaattcaaa attttcgggt ttattacagg gacagcagag atccagtttg gttagtaccg
     2641 ggcccgctct agccctgcag gaactgttta aactgtgagg gcctatttcc catgattcct
     2701 tcatatttgc atatacgata caaggctgtt agagagataa ttggaattaa tttgactgta
     2761 aacacaaaga tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt
     2821 gcagttttaa aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat
     2881 ttcgatttct tggctttata tatcttgtgg aaaggaccaa acaccgccgt ctccttcact
     2941 taactggttt tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga
     3001 aaaagtggca ccgagtcggt gcttttttgt ttggcgcgcc tatggcttaa ttaacatggc
     3061 ggccgcattc taccgggtag gggaggcgct tttcccaagg cagtctggag catgcgcttt
     3121 agcagccccg ctgggcactt ggcgctacac aagtggcctc tggcctcgca cacattccac
     3181 atccaccggt aggcgccaac cggctccgtt ctttggtggc cccttcgcgc caccttctac
     3241 tcctccccta gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca ggacgtgaca
     3301 aatggaagta gcacgtctca ctagtctcgt gcagatggac agcaccgctg agcaatggaa
     3361 gcgggtaggc ctttggggca gcggccaata gcagctttgc tccttcgctt tctgggctca
     3421 gaggctggga aggggtgggt ccgggggcgg gctcaggggc gggctcaggg gcggggcggg
     3481 cgcccgaagg tcctccggag gcccggcatt ctgcacgctt caaaagcgca cgtctgccgc
     3541 gctgttctcc tcttcctcat ctccgggacc cgggatgccc aagaagaaga ggaaggtgtc
     3601 caatttactg accgtacacc aaaatttgcc tgcattaccg gtcgatgcaa cgagtgatga
     3661 ggttcgcaag aacctgatgg acatgttcag ggatcgccag gcgttttctg agcatacctg
     3721 gaaaatgctt ctgtccgttt gccggtcgtg ggcggcatgg tgcaagttga ataaccggaa
     3781 atggtttccc gcagaacctg aagatgttcg cgattatctt ctatatcttc aggcgcgcgg
     3841 tctggcagta aaaactatcc agcaacattt gggccagcta aacatgcttc atcgtcggtc
     3901 cgggctgcca cgaccaagtg acagcaatgc tgtttcactg gttatgcggc ggatccgaaa
     3961 agaaaacgtt gatgccggtg aacgtgcaaa acaggctcta gcgttcgaac gcactgattt
     4021 cgaccaggtt cgttcactca tggaaaatag cgatcgctgc caggatatac gtaatctggc
     4081 atttctgggg attgcttata acaccctgtt acgtatagcc gaaattgcca ggatcagggt
     4141 taaagatatc tcacgtactg acggtgggag aatgttaatc catattggca gaacgaaaac
     4201 gctggttagc accgcaggtg tagagaaggc acttagcctg ggggtaacta aactggtcga
     4261 gcgatggatt tccgtctctg gtgtagctga tgatccgaat aactacctgt tttgccgggt
     4321 cagaaaaaat ggtgttgccg cgccatctgc caccagccag ctatcaactc gcgccctgga
     4381 agggattttt gaagcaactc atcgattgat ttacggcgct aaggatgact ctggtcagag
     4441 atacctggcc tggtctggac acagtgcccg tgtcggagcc gcgcgagata tggcccgcgc
     4501 tggagtttca ataccggaga tcatgcaagc tggtggctgg accaatgtaa atattgtcat
     4561 gaactatatc cgtaacctgg atagtgaaac aggggcaatg gtgcgcctgc tggaagatgg
     4621 cgattagcca tgtttaaggg ttccggttcc actaggtaca attcgatatc aagcttatcg
     4681 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     4741 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     4801 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
     4861 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
     4921 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
     4981 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
     5041 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
     5101 tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
     5161 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
     5221 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc
     5281 gtcgacctcg atcgagacct agaaaaacat ggagcaatca caagtagcaa tacagcagct
     5341 accaatgctg attgtgcctg gctagaagca caagaggagg aggaggtggg ttttccagtc
     5401 acacctcagg tacctttaag accaatgact tacaaggcag ctgtagatct tagccacttt
     5461 ttaaaagaaa aggggggact ggaagggcta attcactccc aacgaagaca agatatcctt
     5521 gatctgtgga tctaccacac acaaggctac ttccctgatt ggcagaacta cacaccaggg
     5581 ccagggatca gatatccact gacctttgga tggtgctaca agctagtacc agttgagcaa
     5641 gagaaggtag aagaagccaa tgaaggagag aacacccgct tgttacaccc tgtgagcctg
     5701 catgggatgg atgacccgga gagagaagta ttagagtgga ggtttgacag ccgcctagca
     5761 tttcatcaca tggcccgaga gctgcatccg gactgtactg ggtctctctg gttagaccag
     5821 atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc
     5881 ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga
     5941 tccctcagac ccttttagtc agtgtggaaa atctctagca gcatgtgagc aaaaggccag
     6001 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     6061 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     6121 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     6181 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     6241 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     6301 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     6361 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     6421 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     6481 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     6541 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     6601 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     6661 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     6721 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     6781 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     6841 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6901 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6961 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     7021 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     7081 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     7141 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     7201 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     7261 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     7321 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     7381 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     7441 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     7501 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     7561 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     7621 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     7681 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     7741 aaaaataaac aaataggggt
//
