LOCUS       sequence-434051-        8302 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector, entry vector for Lentiviral Switch-OVER.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8302)
  AUTHORS   Chylinski K, Hubmann M, Hanna RE, Yanchus C, Michlits G, Uijttewaal 
            ECH, Doench J, Schramek D, Elling U
  TITLE     CRISPR-Switch regulates sgRNA activity by Cre recombination for 
            sequential editing of two loci.
  JOURNAL   Nat Commun. 2019 Nov 29;10(1):5454. doi: 10.1038/s41467-019-13403-y.
  PUBMED    31784531
REFERENCE   2  (bases 1 to 8302)
  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: "Nat 
            Commun."; date: "2019-11-29"; volume: "10(1):5454. doi"; pages: " 
            10.1038/s41467-019-13403-y"
COMMENT     Sequence Label: p1071-Lenti-Single loxP
FEATURES             Location/Qualifiers
     source          1..8302
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(72..93)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     284..303
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    436..660
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     649..666
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     LTR             692..872
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     1377..1393
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1378..1394
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1397..1985
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1503..1514
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1912..1932
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    2060..2093
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        complement(2139..2387)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(2197..2216)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    2422..2539
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     CDS             complement(2837..2881)
                     /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    3066..3299
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    3792..3917
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             3964..4144
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(4162..4360)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4187..4207)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4362..4741
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     4913..4932
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5007..5111
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5112..5180,5181..5972)
                     /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     complement(5330..5349)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6143..6731
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6632..6651
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6885..6902
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7019..7040
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7055..7060,7061..7078,7079..7085)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7093..7109
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     7098..7120
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7117..7133
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7117..7133
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    7182..7303
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7193..7212)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     7247..7266
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(7433..7807)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     promoter        complement(7826..7873)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(7921..8250)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7935..8070
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(7989..8008)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     8235..8255
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(join(8264..8302,1..390))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
       61 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
      121 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
      181 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
      241 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
      301 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
      361 ccgcgcttaa tgcgccgcta cagggcgcgt ggggataccc cctagagccc cagctggttc
      421 tttccgcctc agaagccata gagcccaccg catccccagc atgcctgcta ttgtcttccc
      481 aatcctcccc cttgctgtcc tgccccaccc caccccccag aatagaatga cacctactca
      541 gacaatgcga tgcaatttcc tcattttatt aggaaaggac agtgggagtg gcaccttcca
      601 gggtcaagga aggcacgggg gaggggcaaa caacagatgg ctggcaacta gaaggcacag
      661 tcgaggctga tcagcgggtt taaacgggcc ctgctagaga ttttccacac tgactaaaag
      721 ggtctgaggg atctctagtt accagagtca cacaacagac gggcacacac tacttgaagc
      781 actcaaggca agctttattg aggcttaagc agtgggttcc ctagttagcc agagagctcc
      841 caggctcaga tctggtctaa ccagagagac ccagtacagt ccggatgcag ctctcgggcc
      901 atgtgatgaa atgctaggcg gctgtcaaac ctccactcta atacttctct ctccgggtca
      961 tccatcccat gcaggctcac agggtgtaac aagcgggtgt tctctccttc attggcttct
     1021 tctaccttct cttgctcaac tggtactagc ttgtagcacc atccaaaggt cagtggatat
     1081 ctgatccctg gccctggtgt gtagttctgc caatcaggga agtagccttg tgtgtggtag
     1141 atccacagat caaggatatc ttgtcttcgt tgggagtgaa ttagcccttc cagtcccccc
     1201 ttttctttta aaaagtggct aagatctaca gctgccttgt aagtcattgg tcttaaaggt
     1261 acctgaggtg tgactggaaa acccacctcc tcctcctctt gtgcttctag ccaggcacaa
     1321 tcagcattgg tagctgctgt attgctactt gtgattgctc catgtttttc taggtctcga
     1381 ggtcgacggt atcgatgcgg ggaggcggcc caaagggaga tccgactcgt ctgagggcga
     1441 aggcgaagac gcggaagagg ccgcagagcc ggcagcaggc cgcgggaagg aaggtccgct
     1501 ggattgaggg ccgaagggac gtagcagaag gacgtcccgc gcagaatcca ggtggcaaca
     1561 caggcgagca gccaaggaaa ggacgatgat ttccccgaca acaccacgga attgtcagtg
     1621 cccaacagcc gagcccctgt ccagcagcgg gcaaggcagg cggcgatgag ttccgccgtg
     1681 gcaataggga gggggaaagc gaaagtcccg gaaaggagct gacaggtggt ggcaatgccc
     1741 caaccagtgg gggttgcgtc agcaaacaca gtgcacacca cgccacgttg cctgacaacg
     1801 ggccacaact cctcataaag agacagcaac caggatttat acaaggagga gaaaatgaaa
     1861 gccatacggg aagcaatagc atgatacaaa ggcattaaag cagcgtatcc acatagcgta
     1921 aaaggagcaa catagttaag aataccagtc aatctttcac aaattttgta atccagaggt
     1981 tgattagcaa aaaagcaccg actcggtgcc actttttcaa gttgataacg gactagcctt
     2041 atttaaactt gctatgctga taacttcgta taatgtatgc tatacgaagt tatcagcata
     2101 gctcttaaac agagacgtta cgtatactga tcgtctccgg tgtttcgtcc tttccacaag
     2161 atatataaag ccaagaaatc gaaatacttt caagttacgg taagcatatg atagtccatt
     2221 ttaaaacata attttaaaac tgcaaactac ccaagaaatt attactttct acgtcacgta
     2281 ttttgtacta atatctttgt gtttacagtc aaattaattc taattatctc tctaacagcc
     2341 ttgtatcgta tatgcaaata tgaaggaatc atgggaaata attaaccaaa ctggatctct
     2401 gctgtccctg taataaaccc gaaaattttg aatttttgta atttgttttt gtaattcttt
     2461 agtttgtatg tctgttgcta ttatgtctac tattctttcc cctgcactgt accccccaat
     2521 cccccctttt cttttaaaat tgtggatgaa tactgccatt tgtctgcaga attggcgcac
     2581 gcagtgccga tccgttcact aatcgaatgg atctgtctct gtctctctct ccaccttctt
     2641 cttctattcc ttcgggcctg tcgggtcccc tcggggttgg gaggtgggtc tgaaacgata
     2701 atggtgaata tccctgccta actctattca ctatagaaag tacagcaaaa actattctta
     2761 aacctaccaa gcctcctact atcattatga ataattttat ataccacagc caatttgtta
     2821 tgttaaacca attccacaaa cttgcccatt tatctaattc caataattct tgttcattct
     2881 tttcttgctg gttttgcgat tcttcaatta aggagtgtat taagcttgtg taattgttaa
     2941 tttctctgtc ccactccatc caggtcgtgt gattccaaat ctgttccaga gatttattac
     3001 tccaactagc attccaaggc acagcagtgg tgcaaatgag ttttccagag caaccccaaa
     3061 tccccaggag ctgttgatcc tttaggtatc tttccacagc caggattctt gcctggagct
     3121 gcttgatgcc ccagactgtg agttgcaaca gatgctgttg cgcctcaata gccctcagca
     3181 aattgttctg ctgctgcact ataccagaca ataattgtct ggcctgtacc gtcagcgtca
     3241 ttgacgctgc gcccatagtg cttcctgctg ctcccaagaa cccaaggaac aaagctccta
     3301 ttcccactgc tcttttttct ctctgcacca ctcttctctt tgccttggtg ggtgctactc
     3361 ctaatggttc aatttttact actttatatt tatataattc acttctccaa ttgtccctca
     3421 tatctcctcc tccaggtctg aagatcagcg gccgcttgct gtgcggtggt cttacttttg
     3481 ttttgctctt cctctatctt gtctaaagct tccttggtgt cttttatctc tatcctttga
     3541 tgcacacaat agagggttgc tactgtatta tataatgatc taagttcttc tgatcctgtc
     3601 tgaagggatg gttgtagctg tcccagtatt tgtctacagc cttctgatgt ttctaacagg
     3661 ccaggattaa ctgcgaatcg ttctagctcc ctgcttgccc atactatatg ttttaattta
     3721 tattttttct ttccccctgg ccttaaccga attttttccc atcgcgatct aattctcccc
     3781 cgcttaatac tgacgctctc gcacccatct ctctccttct agcctccgct agtcaaaatt
     3841 tttggcgtac tcaccagtcg ccgcccctcg cctcttgccg tgcgcgcttc agcaagccga
     3901 gtcctgcgtc gagagagctc ctctggtttc cctttcgctt tcaagtccct gttcgggcgc
     3961 cactgctaga gattttccac actgactaaa agggtctgag ggatctctag ttaccagagt
     4021 cacacaacag acgggcacac actacttgaa gcactcaagg caagctttat tgaggcttaa
     4081 gcagtgggtt ccctagttag ccagagagct cccaggctca gatctggtct aaccagagag
     4141 acccagtaca ggcaaaacgc gctgcttata tagacctccc accgtacacg cctaccgccc
     4201 atttgcgtca atggggcgga gttgttacga cattttggaa agtcccgttg attttggtgc
     4261 caaaacaaac tcccattgac gtcaatgggg tggagacttg gaaatccccg tgagtcaaac
     4321 cgctatccac gcccattgat gtactgccaa aaccgcatca ccatggtaat agcgatgact
     4381 aatacgtaga tgtactgcca agtaggaaag tcccataagg tcatgtactg ggcataatgc
     4441 caggcgggcc atttaccgtc attgacgtca atagggggcg tacttggcat atgatacact
     4501 tgatgtactg ccaagtgggc agtttaccgt aaatactcca cccattgacg tcaatggaaa
     4561 gtccctattg gcgttactat gggaacatac gtcattattg acgtcaatgg gcgggggtcg
     4621 ttgggcggtc agccaggcgg gccatttacc gtaagttatg taacgcggaa ctccatatat
     4681 gggctatgaa ctaatgaccc cgtaattgat tactattaat aactagtcaa taatcaatgt
     4741 caacgcgtat atctggcccg tacatcgcga agcagcgcaa aacgcctaac cctaagcaga
     4801 ttcttcatgc aattgtcggt caagccttgc cttgttgtag cttaaatttt gctcgcgcac
     4861 tactcagcga cctccaacac acaagcaggg agcagatact ggcttaacta tgcggcatca
     4921 gagcagattg tactgagagt gcaccatagg ggatcgggag atctcccgat ccgtcgacgt
     4981 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     5041 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     5101 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     5161 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     5221 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     5281 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     5341 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     5401 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     5461 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     5521 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     5581 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     5641 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     5701 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     5761 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     5821 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     5881 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     5941 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     6001 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     6061 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     6121 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     6181 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     6241 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     6301 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     6361 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     6421 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     6481 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     6541 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     6601 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     6661 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     6721 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     6781 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     6841 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     6901 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     6961 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     7021 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     7081 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
     7141 acgccaagct ctagctagag gtcgacggta tacagacatg ataagataca ttgatgagtt
     7201 tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc
     7261 tattgcttta tttgtaacca ttataagctg caataaacaa gttggggtgg gcgaagaact
     7321 ccagcatgag atccccgcgc tggaggatca tccagccggc gtcccggaaa acgattccga
     7381 agcccaacct ttcatagaag gcggcggtgg aatcgaaatc tcgtagcacg tgtcagtcct
     7441 gctcctcggc cacgaagtgc acgcagttgc cggccgggtc gcgcagggcg aactcccgcc
     7501 cccacggctg ctcgccgatc tcggtcatgg ccggcccgga ggcgtcccgg aagttcgtgg
     7561 acacgacctc cgaccactcg gcgtacagct cgtccaggcc gcgcacccac acccaggcca
     7621 gggtgttgtc cggcaccacc tggtcctgga ccgcgctgat gaacagggtc acgtcgtccc
     7681 ggaccacacc ggcgaagtcg tcctccacga agtcccggga gaacccgagc cggtcggtcc
     7741 agaactcgac cgctccggcg acgtcgcgcg cggtgagcac cggaacggca ctggtcaact
     7801 tggccatggt ttagttcctc accttgtcgt attatactat gccgatatac tatgccgatg
     7861 attaattgtc aacacgtgct gatcagatcc gaaaatggat atacaagctc ccgggagctt
     7921 tttgcaaaag cctaggcctc caaaaaagcc tcctcactac ttctggaata gctcagaggc
     7981 agaggcggcc tcggcctctg cataaataaa aaaaattagt cagccatggg gcggagaatg
     8041 ggcggaactg ggcggagtta ggggcgggat gggcggagtt aggggcggga ctatggttgc
     8101 tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc
     8161 acacctggtt gctgactaat tgagatgcat gctttgcata cttctgcctg ctggggagcc
     8221 tggggacttt ccacacccta actgacacac attccacaga attaattcgc gttaaatttt
     8281 tgttaaatca gctcattttt ta
//
