LOCUS       sequence-363585-        7760 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a Lrp1b-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-sgLrp1b#1/Cre
FEATURES             Location/Qualifiers
     source          1..7760
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(105..124)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        296..675
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        677..875
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     830..850
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             893..1073
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1120..1245
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1742..1975
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2160..2204
                     /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    2502..2619
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2701..2949
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2701..2721
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2872..2891
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2971..3046
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3099..3598
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3134..3157)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3531..3550
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3602..3622
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3623..4651
                     /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    4707..5295
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4760..4780)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5178..5189)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5298..5314)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5299..5315)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5824..6004
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6066..6654)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6146..6165)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6825..7616,7617..7685))
                     /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     7448..7467
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7686..7760,1..30))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
       61 cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat
      121 gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc
      181 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
      241 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat
      301 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
      361 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
      421 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
      481 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      541 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      601 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      661 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
      721 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
      781 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
      841 ggtaggcgtg tacggtggga ggtctatata agcagcgcgt tttgcctgta ctgggtctct
      901 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
      961 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
     1021 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc
     1081 ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg caggactcgg
     1141 cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac gccaaaaatt
     1201 ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat taagcggggg
     1261 agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa
     1321 ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa tcctggcctg
     1381 ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc ccttcagaca
     1441 ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg tgtgcatcaa
     1501 aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga gcaaaacaaa
     1561 agtaagacca ccgcacagca agcggccggc cgctgatctt cagacctgga ggaggagata
     1621 tgagggacaa ttggagaagt gaattatata aatataaagt agtaaaaatt gaaccattag
     1681 gagtagcacc caccaaggca aagagaagag tggtgcagag agaaaaaaga gcagtgggaa
     1741 taggagcttt gttccttggg ttcttgggag cagcaggaag cactatgggc gcagcgtcaa
     1801 tgacgctgac ggtacaggcc agacaattat tgtctggtat agtgcagcag cagaacaatt
     1861 tgctgagggc tattgaggcg caacagcatc tgttgcaact cacagtctgg ggcatcaagc
     1921 agctccaggc aagaatcctg gctgtggaaa gatacctaaa ggatcaacag ctcctgggga
     1981 tttggggttg ctctggaaaa ctcatttgca ccactgctgt gccttggaat gctagttgga
     2041 gtaataaatc tctggaacag atttggaatc acacgacctg gatggagtgg gacagagaaa
     2101 ttaacaatta cacaagctta atacactcct taattgaaga atcgcaaaac cagcaagaaa
     2161 agaatgaaca agaattattg gaattagata aatgggcaag tttgtggaat tggtttaaca
     2221 taacaaattg gctgtggtat ataaaattat tcataatgat agtaggaggc ttggtaggtt
     2281 taagaatagt ttttgctgta ctttctatag tgaatagagt taggcaggga tattcaccat
     2341 tatcgtttca gacccacctc ccaaccccga ggggacccga caggcccgaa ggaatagaag
     2401 aagaaggtgg agagagagac agagacagat ccattcgatt agtgaacgga tcggcactgc
     2461 gtgcgccaat tctgcagaca aatggcagta ttcatccaca attttaaaag aaaagggggg
     2521 attggggggt acagtgcagg ggaaagaata gtagacataa tagcaacaga catacaaact
     2581 aaagaattac aaaaacaaat tacaaaaatt caaaattttc gggtttatta cagggacagc
     2641 agagatccag tttggttagt accgggcccg ctctagccct gcaggaactg tttaaactgt
     2701 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
     2761 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
     2821 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
     2881 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
     2941 ccaaacaccg aagacgttct gagatccgag gttttagagc tagaaatagc aagttaaaat
     3001 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgtttggcg
     3061 cgcctatggc ttaattaaca tggcggccgc attctaccgg gtaggggagg cgcttttccc
     3121 aaggcagtct ggagcatgcg ctttagcagc cccgctgggc acttggcgct acacaagtgg
     3181 cctctggcct cgcacacatt ccacatccac cggtaggcgc caaccggctc cgttctttgg
     3241 tggccccttc gcgccacctt ctactcctcc cctagtcagg aagttccccc ccgccccgca
     3301 gctcgcgtcg tgcaggacgt gacaaatgga agtagcacgt ctcactagtc tcgtgcagat
     3361 ggacagcacc gctgagcaat ggaagcgggt aggcctttgg ggcagcggcc aatagcagct
     3421 ttgctccttc gctttctggg ctcagaggct gggaaggggt gggtccgggg gcgggctcag
     3481 gggcgggctc aggggcgggg cgggcgcccg aaggtcctcc ggaggcccgg cattctgcac
     3541 gcttcaaaag cgcacgtctg ccgcgctgtt ctcctcttcc tcatctccgg gacccgggat
     3601 gcccaagaag aagaggaagg tgtccaattt actgaccgta caccaaaatt tgcctgcatt
     3661 accggtcgat gcaacgagtg atgaggttcg caagaacctg atggacatgt tcagggatcg
     3721 ccaggcgttt tctgagcata cctggaaaat gcttctgtcc gtttgccggt cgtgggcggc
     3781 atggtgcaag ttgaataacc ggaaatggtt tcccgcagaa cctgaagatg ttcgcgatta
     3841 tcttctatat cttcaggcgc gcggtctggc agtaaaaact atccagcaac atttgggcca
     3901 gctaaacatg cttcatcgtc ggtccgggct gccacgacca agtgacagca atgctgtttc
     3961 actggttatg cggcggatcc gaaaagaaaa cgttgatgcc ggtgaacgtg caaaacaggc
     4021 tctagcgttc gaacgcactg atttcgacca ggttcgttca ctcatggaaa atagcgatcg
     4081 ctgccaggat atacgtaatc tggcatttct ggggattgct tataacaccc tgttacgtat
     4141 agccgaaatt gccaggatca gggttaaaga tatctcacgt actgacggtg ggagaatgtt
     4201 aatccatatt ggcagaacga aaacgctggt tagcaccgca ggtgtagaga aggcacttag
     4261 cctgggggta actaaactgg tcgagcgatg gatttccgtc tctggtgtag ctgatgatcc
     4321 gaataactac ctgttttgcc gggtcagaaa aaatggtgtt gccgcgccat ctgccaccag
     4381 ccagctatca actcgcgccc tggaagggat ttttgaagca actcatcgat tgatttacgg
     4441 cgctaaggat gactctggtc agagatacct ggcctggtct ggacacagtg cccgtgtcgg
     4501 agccgcgcga gatatggccc gcgctggagt ttcaataccg gagatcatgc aagctggtgg
     4561 ctggaccaat gtaaatattg tcatgaacta tatccgtaac ctggatagtg aaacaggggc
     4621 aatggtgcgc ctgctggaag atggcgatta gccatgttta agggttccgg ttccactagg
     4681 tacaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat
     4741 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     4801 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     4861 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     4921 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     4981 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     5041 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     5101 aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt
     5161 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     5221 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     5281 gggccgcctc cccgcatcga taccgtcgac ctcgatcgag acctagaaaa acatggagca
     5341 atcacaagta gcaatacagc agctaccaat gctgattgtg cctggctaga agcacaagag
     5401 gaggaggagg tgggttttcc agtcacacct caggtacctt taagaccaat gacttacaag
     5461 gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg gctaattcac
     5521 tcccaacgaa gacaagatat ccttgatctg tggatctacc acacacaagg ctacttccct
     5581 gattggcaga actacacacc agggccaggg atcagatatc cactgacctt tggatggtgc
     5641 tacaagctag taccagttga gcaagagaag gtagaagaag ccaatgaagg agagaacacc
     5701 cgcttgttac accctgtgag cctgcatggg atggatgacc cggagagaga agtattagag
     5761 tggaggtttg acagccgcct agcatttcat cacatggccc gagagctgca tccggactgt
     5821 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
     5881 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
     5941 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
     6001 agcagcatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     6061 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     6121 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     6181 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     6241 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     6301 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     6361 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     6421 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     6481 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     6541 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     6601 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     6661 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     6721 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     6781 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     6841 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     6901 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     6961 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     7021 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     7081 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     7141 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     7201 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     7261 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     7321 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     7381 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     7441 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     7501 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     7561 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     7621 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     7681 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     7741 ggatacatat ttgaatgtat
//
