LOCUS       sequence-379586-        7770 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Lentiviral vector expressing Cre recombinase (driven by mPGK 
            promoter) and an sgRNA against the mouse Spen gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7770)
  AUTHORS   Lee MC, Cai H, Murray CW, Li C, Shue YT, Andrejka L, He AL, Holzem 
            AME, Drainas AP, Ko JH, Coles GL, Kong C, Zhu S, Zhu C, Wang J, van 
            de Rijn M, Petrov DA, Winslow MM, Sage J
  TITLE     A multiplexed in vivo approach to identify driver genes in small 
            cell lung cancer.
  JOURNAL   Cell Rep. 2023 Jan 13;42(1):111990. doi: 
            10.1016/j.celrep.2023.111990.
  PUBMED    36640300
REFERENCE   2  (bases 1 to 7770)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell Rep.";
            date: "2023-01-13"; volume: "42(1):111990. doi"; pages: " 
            10.1016/j.celrep.2023.111990"
COMMENT     Sequence Label: Lenti-sgSpen#1/Cre
FEATURES             Location/Qualifiers
     source          1..7770
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..580
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             598..778
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    825..950
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1447..1680
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1865..1909
                     /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    2207..2324
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2406..2654
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2406..2426
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2577..2596
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        2814..3313
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2849..2872)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3246..3265
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3317..3337
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3338..4366
                     /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    4422..5010
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4475..4495)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4893..4904)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5013..5029)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5014..5030)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5539..5719
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5781..6369)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5861..5880)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6540..7331,7332..7400))
                     /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     7163..7182
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7401..7505)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7580..7599)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg
      601 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
      661 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
      721 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
      781 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga
      841 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa
      901 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc
      961 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat
     1021 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg
     1081 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc
     1141 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc
     1201 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa
     1261 acaaaagtaa gaccaccgca cagcaagcgg ccggccgctg atcttcagac ctggaggagg
     1321 agatatgagg gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc
     1381 attaggagta gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt
     1441 gggaatagga gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc
     1501 gtcaatgacg ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa
     1561 caatttgctg agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat
     1621 caagcagctc caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct
     1681 ggggatttgg ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag
     1741 ttggagtaat aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag
     1801 agaaattaac aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca
     1861 agaaaagaat gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt
     1921 taacataaca aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt
     1981 aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc
     2041 accattatcg tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat
     2101 agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga acggatcggc
     2161 actgcgtgcg ccaattctgc agacaaatgg cagtattcat ccacaatttt aaaagaaaag
     2221 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2281 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2341 acagcagaga tccagtttgg ttagtaccgg gcccgctcta gccctgcagg aactgtttaa
     2401 actgtgaggg cctatttccc atgattcctt catatttgca tatacgatac aaggctgtta
     2461 gagagataat tggaattaat ttgactgtaa acacaaagat attagtacaa aatacgtgac
     2521 gtagaaagta ataatttctt gggtagtttg cagttttaaa attatgtttt aaaatggact
     2581 atcatatgct taccgtaact tgaaagtatt tcgatttctt ggctttatat atcttgtgga
     2641 aaggaccaaa caccgaagag gtccagctct cctcggttta agagctatgc tggaaacagc
     2701 atagcaagtt taaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg
     2761 cttttttgtt tggcgcgcct atggcttaat taacatggcg gccgcattct accgggtagg
     2821 ggaggcgctt ttcccaaggc agtctggagc atgcgcttta gcagccccgc tgggcacttg
     2881 gcgctacaca agtggcctct ggcctcgcac acattccaca tccaccggta ggcgccaacc
     2941 ggctccgttc tttggtggcc ccttcgcgcc accttctact cctcccctag tcaggaagtt
     3001 cccccccgcc ccgcagctcg cgtcgtgcag gacgtgacaa atggaagtag cacgtctcac
     3061 tagtctcgtg cagatggaca gcaccgctga gcaatggaag cgggtaggcc tttggggcag
     3121 cggccaatag cagctttgct ccttcgcttt ctgggctcag aggctgggaa ggggtgggtc
     3181 cgggggcggg ctcaggggcg ggctcagggg cggggcgggc gcccgaaggt cctccggagg
     3241 cccggcattc tgcacgcttc aaaagcgcac gtctgccgcg ctgttctcct cttcctcatc
     3301 tccgggaccc gggatgccca agaagaagag gaaggtgtcc aatttactga ccgtacacca
     3361 aaatttgcct gcattaccgg tcgatgcaac gagtgatgag gttcgcaaga acctgatgga
     3421 catgttcagg gatcgccagg cgttttctga gcatacctgg aaaatgcttc tgtccgtttg
     3481 ccggtcgtgg gcggcatggt gcaagttgaa taaccggaaa tggtttcccg cagaacctga
     3541 agatgttcgc gattatcttc tatatcttca ggcgcgcggt ctggcagtaa aaactatcca
     3601 gcaacatttg ggccagctaa acatgcttca tcgtcggtcc gggctgccac gaccaagtga
     3661 cagcaatgct gtttcactgg ttatgcggcg gatccgaaaa gaaaacgttg atgccggtga
     3721 acgtgcaaaa caggctctag cgttcgaacg cactgatttc gaccaggttc gttcactcat
     3781 ggaaaatagc gatcgctgcc aggatatacg taatctggca tttctgggga ttgcttataa
     3841 caccctgtta cgtatagccg aaattgccag gatcagggtt aaagatatct cacgtactga
     3901 cggtgggaga atgttaatcc atattggcag aacgaaaacg ctggttagca ccgcaggtgt
     3961 agagaaggca cttagcctgg gggtaactaa actggtcgag cgatggattt ccgtctctgg
     4021 tgtagctgat gatccgaata actacctgtt ttgccgggtc agaaaaaatg gtgttgccgc
     4081 gccatctgcc accagccagc tatcaactcg cgccctggaa gggatttttg aagcaactca
     4141 tcgattgatt tacggcgcta aggatgactc tggtcagaga tacctggcct ggtctggaca
     4201 cagtgcccgt gtcggagccg cgcgagatat ggcccgcgct ggagtttcaa taccggagat
     4261 catgcaagct ggtggctgga ccaatgtaaa tattgtcatg aactatatcc gtaacctgga
     4321 tagtgaaaca ggggcaatgg tgcgcctgct ggaagatggc gattagccat gtttaagggt
     4381 tccggttcca ctaggtacaa ttcgatatca agcttatcga taatcaacct ctggattaca
     4441 aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat
     4501 acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct
     4561 ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac
     4621 gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca
     4681 cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca
     4741 tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg
     4801 tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga
     4861 ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt
     4921 cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga
     4981 gtcggatctc cctttgggcc gcctccccgc atcgataccg tcgacctcga tcgagaccta
     5041 gaaaaacatg gagcaatcac aagtagcaat acagcagcta ccaatgctga ttgtgcctgg
     5101 ctagaagcac aagaggagga ggaggtgggt tttccagtca cacctcaggt acctttaaga
     5161 ccaatgactt acaaggcagc tgtagatctt agccactttt taaaagaaaa ggggggactg
     5221 gaagggctaa ttcactccca acgaagacaa gatatccttg atctgtggat ctaccacaca
     5281 caaggctact tccctgattg gcagaactac acaccagggc cagggatcag atatccactg
     5341 acctttggat ggtgctacaa gctagtacca gttgagcaag agaaggtaga agaagccaat
     5401 gaaggagaga acacccgctt gttacaccct gtgagcctgc atgggatgga tgacccggag
     5461 agagaagtat tagagtggag gtttgacagc cgcctagcat ttcatcacat ggcccgagag
     5521 ctgcatccgg actgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct
     5581 ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta
     5641 gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca
     5701 gtgtggaaaa tctctagcag catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     5761 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     5821 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     5881 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     5941 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     6001 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     6061 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     6121 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     6181 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     6241 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     6301 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     6361 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     6421 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     6481 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     6541 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6601 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6661 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6721 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6781 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     6841 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     6901 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     6961 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     7021 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     7081 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     7141 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     7201 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     7261 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     7321 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     7381 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     7441 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     7501 ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc
     7561 ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc
     7621 tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa
     7681 ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc
     7741 ttcgcgatgt acgggccaga tatacgcgtt
//
