LOCUS       sequence-363562-        7760 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a Gata3-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-sgGata3#1/Cre
FEATURES             Location/Qualifiers
     source          1..7760
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     57..76
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             128..148
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             149..1177
                     /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    1233..1821
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1286..1306)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1704..1715)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(1824..1840)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1825..1841)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             2350..2530
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(2592..3180)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2672..2691)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3351..4142,4143..4211))
                     /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     3974..3993
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4212..4316)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4391..4410)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4582..4961
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4963..5161
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5116..5136
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             5179..5359
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5406..5531
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6028..6261
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6446..6490
                     /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    6788..6905
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        6987..7235
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6987..7007
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     7158..7177
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        7257..7332
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        join(7385..7760,1..124)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(7420..7443)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
ORIGIN
        1 gctcaggggc gggctcaggg gcggggcggg cgcccgaagg tcctccggag gcccggcatt
       61 ctgcacgctt caaaagcgca cgtctgccgc gctgttctcc tcttcctcat ctccgggacc
      121 cgggatgccc aagaagaaga ggaaggtgtc caatttactg accgtacacc aaaatttgcc
      181 tgcattaccg gtcgatgcaa cgagtgatga ggttcgcaag aacctgatgg acatgttcag
      241 ggatcgccag gcgttttctg agcatacctg gaaaatgctt ctgtccgttt gccggtcgtg
      301 ggcggcatgg tgcaagttga ataaccggaa atggtttccc gcagaacctg aagatgttcg
      361 cgattatctt ctatatcttc aggcgcgcgg tctggcagta aaaactatcc agcaacattt
      421 gggccagcta aacatgcttc atcgtcggtc cgggctgcca cgaccaagtg acagcaatgc
      481 tgtttcactg gttatgcggc ggatccgaaa agaaaacgtt gatgccggtg aacgtgcaaa
      541 acaggctcta gcgttcgaac gcactgattt cgaccaggtt cgttcactca tggaaaatag
      601 cgatcgctgc caggatatac gtaatctggc atttctgggg attgcttata acaccctgtt
      661 acgtatagcc gaaattgcca ggatcagggt taaagatatc tcacgtactg acggtgggag
      721 aatgttaatc catattggca gaacgaaaac gctggttagc accgcaggtg tagagaaggc
      781 acttagcctg ggggtaacta aactggtcga gcgatggatt tccgtctctg gtgtagctga
      841 tgatccgaat aactacctgt tttgccgggt cagaaaaaat ggtgttgccg cgccatctgc
      901 caccagccag ctatcaactc gcgccctgga agggattttt gaagcaactc atcgattgat
      961 ttacggcgct aaggatgact ctggtcagag atacctggcc tggtctggac acagtgcccg
     1021 tgtcggagcc gcgcgagata tggcccgcgc tggagtttca ataccggaga tcatgcaagc
     1081 tggtggctgg accaatgtaa atattgtcat gaactatatc cgtaacctgg atagtgaaac
     1141 aggggcaatg gtgcgcctgc tggaagatgg cgattagcca tgtttaaggg ttccggttcc
     1201 actaggtaca attcgatatc aagcttatcg ataatcaacc tctggattac aaaatttgtg
     1261 aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt
     1321 taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata
     1381 aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg
     1441 tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc
     1501 tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct
     1561 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt
     1621 cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg
     1681 ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc
     1741 tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct
     1801 ccctttgggc cgcctccccg catcgatacc gtcgacctcg atcgagacct agaaaaacat
     1861 ggagcaatca caagtagcaa tacagcagct accaatgctg attgtgcctg gctagaagca
     1921 caagaggagg aggaggtggg ttttccagtc acacctcagg tacctttaag accaatgact
     1981 tacaaggcag ctgtagatct tagccacttt ttaaaagaaa aggggggact ggaagggcta
     2041 attcactccc aacgaagaca agatatcctt gatctgtgga tctaccacac acaaggctac
     2101 ttccctgatt ggcagaacta cacaccaggg ccagggatca gatatccact gacctttgga
     2161 tggtgctaca agctagtacc agttgagcaa gagaaggtag aagaagccaa tgaaggagag
     2221 aacacccgct tgttacaccc tgtgagcctg catgggatgg atgacccgga gagagaagta
     2281 ttagagtgga ggtttgacag ccgcctagca tttcatcaca tggcccgaga gctgcatccg
     2341 gactgtactg ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta
     2401 gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc
     2461 cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa
     2521 atctctagca gcatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     2581 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     2641 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     2701 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     2761 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     2821 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     2881 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     2941 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     3001 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     3061 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     3121 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     3181 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     3241 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     3301 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     3361 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     3421 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     3481 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     3541 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     3601 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     3661 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     3721 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     3781 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     3841 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     3901 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     3961 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     4021 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     4081 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     4141 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     4201 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     4261 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     4321 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
     4381 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
     4441 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
     4501 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
     4561 tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta
     4621 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
     4681 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
     4741 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
     4801 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
     4861 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
     4921 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
     4981 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
     5041 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
     5101 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
     5161 gcgcgttttg cctgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct
     5221 ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta
     5281 gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca
     5341 gtgtggaaaa tctctagcag tggcgcccga acagggactt gaaagcgaaa gggaaaccag
     5401 aggagctctc tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc
     5461 ggcgactggt gagtacgcca aaaattttga ctagcggagg ctagaaggag agagatgggt
     5521 gcgagagcgt cagtattaag cgggggagaa ttagatcgcg atgggaaaaa attcggttaa
     5581 ggccaggggg aaagaaaaaa tataaattaa aacatatagt atgggcaagc agggagctag
     5641 aacgattcgc agttaatcct ggcctgttag aaacatcaga aggctgtaga caaatactgg
     5701 gacagctaca accatccctt cagacaggat cagaagaact tagatcatta tataatacag
     5761 tagcaaccct ctattgtgtg catcaaagga tagagataaa agacaccaag gaagctttag
     5821 acaagataga ggaagagcaa aacaaaagta agaccaccgc acagcaagcg gccggccgct
     5881 gatcttcaga cctggaggag gagatatgag ggacaattgg agaagtgaat tatataaata
     5941 taaagtagta aaaattgaac cattaggagt agcacccacc aaggcaaaga gaagagtggt
     6001 gcagagagaa aaaagagcag tgggaatagg agctttgttc cttgggttct tgggagcagc
     6061 aggaagcact atgggcgcag cgtcaatgac gctgacggta caggccagac aattattgtc
     6121 tggtatagtg cagcagcaga acaatttgct gagggctatt gaggcgcaac agcatctgtt
     6181 gcaactcaca gtctggggca tcaagcagct ccaggcaaga atcctggctg tggaaagata
     6241 cctaaaggat caacagctcc tggggatttg gggttgctct ggaaaactca tttgcaccac
     6301 tgctgtgcct tggaatgcta gttggagtaa taaatctctg gaacagattt ggaatcacac
     6361 gacctggatg gagtgggaca gagaaattaa caattacaca agcttaatac actccttaat
     6421 tgaagaatcg caaaaccagc aagaaaagaa tgaacaagaa ttattggaat tagataaatg
     6481 ggcaagtttg tggaattggt ttaacataac aaattggctg tggtatataa aattattcat
     6541 aatgatagta ggaggcttgg taggtttaag aatagttttt gctgtacttt ctatagtgaa
     6601 tagagttagg cagggatatt caccattatc gtttcagacc cacctcccaa ccccgagggg
     6661 acccgacagg cccgaaggaa tagaagaaga aggtggagag agagacagag acagatccat
     6721 tcgattagtg aacggatcgg cactgcgtgc gccaattctg cagacaaatg gcagtattca
     6781 tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag
     6841 acataatagc aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa
     6901 attttcgggt ttattacagg gacagcagag atccagtttg gttagtaccg ggcccgctct
     6961 agccctgcag gaactgttta aactgtgagg gcctatttcc catgattcct tcatatttgc
     7021 atatacgata caaggctgtt agagagataa ttggaattaa tttgactgta aacacaaaga
     7081 tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt gcagttttaa
     7141 aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct
     7201 tggctttata tatcttgtgg aaaggaccaa acaccgctac tacggaaact ccgtcagttt
     7261 tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga aaaagtggca
     7321 ccgagtcggt gcttttttgt ttggcgcgcc tatggcttaa ttaacatggc ggccgcattc
     7381 taccgggtag gggaggcgct tttcccaagg cagtctggag catgcgcttt agcagccccg
     7441 ctgggcactt ggcgctacac aagtggcctc tggcctcgca cacattccac atccaccggt
     7501 aggcgccaac cggctccgtt ctttggtggc cccttcgcgc caccttctac tcctccccta
     7561 gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca ggacgtgaca aatggaagta
     7621 gcacgtctca ctagtctcgt gcagatggac agcaccgctg agcaatggaa gcgggtaggc
     7681 ctttggggca gcggccaata gcagctttgc tccttcgctt tctgggctca gaggctggga
     7741 aggggcgggt ccgggggcgg
//
