LOCUS       sequence-363656-        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#2/Cre
FEATURES             Location/Qualifiers
     source          1..7760
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(110..129)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        301..680
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        682..880
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     835..855
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             898..1078
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1125..1250
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1747..1980
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2165..2209
                     /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    2507..2624
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2706..2954
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2706..2726
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2877..2896
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2976..3051
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3104..3603
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3139..3162)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3536..3555
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3607..3627
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3628..4656
                     /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    4712..5300
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4765..4785)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5183..5194)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5303..5319)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5304..5320)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5829..6009
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6071..6659)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6151..6170)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6830..7621,7622..7690))
                     /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     7453..7472
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7691..7760,1..35))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
       61 gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt acaatctgct
      121 ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt
      181 agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga
      241 atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt
      301 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
      361 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      421 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      481 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      541 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      601 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      661 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      721 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      781 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      841 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg
      901 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
      961 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
     1021 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
     1081 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga
     1141 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa
     1201 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc
     1261 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat
     1321 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg
     1381 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc
     1441 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc
     1501 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa
     1561 acaaaagtaa gaccaccgca cagcaagcgg ccggccgctg atcttcagac ctggaggagg
     1621 agatatgagg gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc
     1681 attaggagta gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt
     1741 gggaatagga gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc
     1801 gtcaatgacg ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa
     1861 caatttgctg agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat
     1921 caagcagctc caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct
     1981 ggggatttgg ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag
     2041 ttggagtaat aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag
     2101 agaaattaac aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca
     2161 agaaaagaat gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt
     2221 taacataaca aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt
     2281 aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc
     2341 accattatcg tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat
     2401 agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga acggatcggc
     2461 actgcgtgcg ccaattctgc agacaaatgg cagtattcat ccacaatttt aaaagaaaag
     2521 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2581 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2641 acagcagaga tccagtttgg ttagtaccgg gcccgctcta gccctgcagg aactgtttaa
     2701 actgtgaggg cctatttccc atgattcctt catatttgca tatacgatac aaggctgtta
     2761 gagagataat tggaattaat ttgactgtaa acacaaagat attagtacaa aatacgtgac
     2821 gtagaaagta ataatttctt gggtagtttg cagttttaaa attatgtttt aaaatggact
     2881 atcatatgct taccgtaact tgaaagtatt tcgatttctt ggctttatat atcttgtgga
     2941 aaggaccaaa caccgtctgg gagacacaag tcacagtttt agagctagaa atagcaagtt
     3001 aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg cttttttgtt
     3061 tggcgcgcct atggcttaat taacatggcg gccgcattct accgggtagg ggaggcgctt
     3121 ttcccaaggc agtctggagc atgcgcttta gcagccccgc tgggcacttg gcgctacaca
     3181 agtggcctct ggcctcgcac acattccaca tccaccggta ggcgccaacc ggctccgttc
     3241 tttggtggcc ccttcgcgcc accttctact cctcccctag tcaggaagtt cccccccgcc
     3301 ccgcagctcg cgtcgtgcag gacgtgacaa atggaagtag cacgtctcac tagtctcgtg
     3361 cagatggaca gcaccgctga gcaatggaag cgggtaggcc tttggggcag cggccaatag
     3421 cagctttgct ccttcgcttt ctgggctcag aggctgggaa ggggtgggtc cgggggcggg
     3481 ctcaggggcg ggctcagggg cggggcgggc gcccgaaggt cctccggagg cccggcattc
     3541 tgcacgcttc aaaagcgcac gtctgccgcg ctgttctcct cttcctcatc tccgggaccc
     3601 gggatgccca agaagaagag gaaggtgtcc aatttactga ccgtacacca aaatttgcct
     3661 gcattaccgg tcgatgcaac gagtgatgag gttcgcaaga acctgatgga catgttcagg
     3721 gatcgccagg cgttttctga gcatacctgg aaaatgcttc tgtccgtttg ccggtcgtgg
     3781 gcggcatggt gcaagttgaa taaccggaaa tggtttcccg cagaacctga agatgttcgc
     3841 gattatcttc tatatcttca ggcgcgcggt ctggcagtaa aaactatcca gcaacatttg
     3901 ggccagctaa acatgcttca tcgtcggtcc gggctgccac gaccaagtga cagcaatgct
     3961 gtttcactgg ttatgcggcg gatccgaaaa gaaaacgttg atgccggtga acgtgcaaaa
     4021 caggctctag cgttcgaacg cactgatttc gaccaggttc gttcactcat ggaaaatagc
     4081 gatcgctgcc aggatatacg taatctggca tttctgggga ttgcttataa caccctgtta
     4141 cgtatagccg aaattgccag gatcagggtt aaagatatct cacgtactga cggtgggaga
     4201 atgttaatcc atattggcag aacgaaaacg ctggttagca ccgcaggtgt agagaaggca
     4261 cttagcctgg gggtaactaa actggtcgag cgatggattt ccgtctctgg tgtagctgat
     4321 gatccgaata actacctgtt ttgccgggtc agaaaaaatg gtgttgccgc gccatctgcc
     4381 accagccagc tatcaactcg cgccctggaa gggatttttg aagcaactca tcgattgatt
     4441 tacggcgcta aggatgactc tggtcagaga tacctggcct ggtctggaca cagtgcccgt
     4501 gtcggagccg cgcgagatat ggcccgcgct ggagtttcaa taccggagat catgcaagct
     4561 ggtggctgga ccaatgtaaa tattgtcatg aactatatcc gtaacctgga tagtgaaaca
     4621 ggggcaatgg tgcgcctgct ggaagatggc gattagccat gtttaagggt tccggttcca
     4681 ctaggtacaa ttcgatatca agcttatcga taatcaacct ctggattaca aaatttgtga
     4741 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     4801 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     4861 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     4921 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     4981 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     5041 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     5101 ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     5161 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     5221 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     5281 cctttgggcc gcctccccgc atcgataccg tcgacctcga tcgagaccta gaaaaacatg
     5341 gagcaatcac aagtagcaat acagcagcta ccaatgctga ttgtgcctgg ctagaagcac
     5401 aagaggagga ggaggtgggt tttccagtca cacctcaggt acctttaaga ccaatgactt
     5461 acaaggcagc tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa
     5521 ttcactccca acgaagacaa gatatccttg atctgtggat ctaccacaca caaggctact
     5581 tccctgattg gcagaactac acaccagggc cagggatcag atatccactg acctttggat
     5641 ggtgctacaa gctagtacca gttgagcaag agaaggtaga agaagccaat gaaggagaga
     5701 acacccgctt gttacaccct gtgagcctgc atgggatgga tgacccggag agagaagtat
     5761 tagagtggag gtttgacagc cgcctagcat ttcatcacat ggcccgagag ctgcatccgg
     5821 actgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     5881 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     5941 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     6001 tctctagcag catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     6061 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     6121 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     6181 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     6241 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     6301 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     6361 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     6421 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     6481 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     6541 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     6601 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     6661 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6721 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     6781 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     6841 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     6901 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     6961 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     7021 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     7081 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     7141 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     7201 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     7261 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     7321 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     7381 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     7441 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     7501 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     7561 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     7621 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     7681 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     7741 tgagcggata catatttgaa
//
