LOCUS       sequence-441210-        8129 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral acceptor vector for pegRNAs with blasticidin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8129)
  AUTHORS   Koeppel J, Weller J, Peets EM, Pallaseni A, Kuzmin I, Raudvere U, 
            Peterson H, Liberante FG, Parts L
  TITLE     Prediction of prime editing insertion efficiencies using sequence 
            features and DNA repair determinants.
  JOURNAL   Nat Biotechnol. 2023 Feb 16. doi: 10.1038/s41587-023-01678-y.
  PUBMED    36797492
REFERENCE   2  (bases 1 to 8129)
  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 
            Biotechnol. 2023 Feb 16. doi: 10.1038/s41587-023-01678-y."
COMMENT     Sequence Label: pLentiGuide-BlastR
FEATURES             Location/Qualifiers
     source          1..8129
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        86..1264
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          317..1255
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1212..1232
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1271..1280
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1277..1675
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    1691..2279
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1744..1764)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2162..2173)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2351..2584
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2662..2783
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2699..2718)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2753..2772
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2823..2958
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2978..2997)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2979..2997)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3007..3024)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3007..3023)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3016..3038)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3165..3620
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3252..3271)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3462..3483
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3646..3750
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3751..3819,3820..4611)
                     /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(3969..3988)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4782..5370
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5271..5290
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5524..5541
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5658..5679
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5694..5699,5700..5717,5718..5724)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5732..5748
                     /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     5737..5759
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5756..5772
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5756..5772
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5791..5811
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5793..5811
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5839..6065
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6066..6246
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6293..6418
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6911..7144
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7329..7373
                     /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"
     promoter        7551..7799
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     7551..7571
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     7722..7741
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    7959..8076
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 ccgggtgcaa agatggataa agttttaaac agagaggaat ctttgcagct aatggacctt
       61 ctaggtcttg aaaggagtgg gaattggctc cggtgcccgt cagtgggcag agcgcacatc
      121 gcccacagtc cccgagaagt tggggggagg ggtcggcaat tgatccggtg cctagagaag
      181 gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg
      241 tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt
      301 tgccgccaga acacaggtaa gtgccgtgtg tggttcccgc gggcctggcc tctttacggg
      361 ttatggccct tgcgtgcctt gaattacttc cacctggctg cagtacgtga ttcttgatcc
      421 cgagcttcgg gttggaagtg ggtgggagag ttcgaggcct tgcgcttaag gagccccttc
      481 gcctcgtgct tgagttgagg cctggcctgg gcgctggggc cgccgcgtgc gaatctggtg
      541 gcaccttcgc gcctgtctcg ctgctttcga taagtctcta gccatttaaa atttttgatg
      601 acctgctgcg acgctttttt tctggcaaga tagtcttgta aatgcgggcc aagatctgca
      661 cactggtatt tcggtttttg gggccgcggg cggcgacggg gcccgtgcgt cccagcgcac
      721 atgttcggcg aggcggggcc tgcgagcgcg gccaccgaga atcggacggg ggtagtctca
      781 agctggccgg cctgctctgg tgcctggcct cgcgccgccg tgtatcgccc cgccctgggc
      841 ggcaaggctg gcccggtcgg caccagttgc gtgagcggaa agatggccgc ttcccggccc
      901 tgctgcaggg agctcaaaat ggaggacgcg gcgctcggga gagcgggcgg gtgagtcacc
      961 cacacaaagg aaaagggcct ttccgtcctc agccgtcgct tcatgtgact ccacggagta
     1021 ccgggcgccg tccaggcacc tcgattagtt ctcgagcttt tggagtacgt cgtctttagg
     1081 ttggggggag gggttttatg cgatggagtt tccccacact gagtgggtgg agactgaagt
     1141 taggccagct tggcacttga tgtaattctc cttggaattt gccctttttg agtttggatc
     1201 ttggttcatt ctcaagcctc agacagtggt tcaaagtttt tttcttccat ttcaggtgtc
     1261 gtgacgtacg gccaccatgg ccaagccttt gtctcaagaa gaatccaccc tcattgaaag
     1321 agcaacggct acaatcaaca gcatccccat ctctgaagac tacagcgtcg ccagcgcagc
     1381 tctctctagc gacggccgca tcttcactgg tgtcaatgta tatcatttta ctgggggacc
     1441 ttgtgcagaa ctcgtggtgc tgggcactgc tgctgctgcg gcagctggca acctgacttg
     1501 tatcgtcgcg atcggaaatg agaacagggg catcttgagc ccctgcggac ggtgccgaca
     1561 ggtgcttctc gatctgcatc ctgggatcaa agccatagtg aaggacagtg atggacagcc
     1621 gacggcagtt gggattcgtg aattgctgcc ctctggttat gtgtgggagg gctaaacgcg
     1681 ttaagtcgac aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
     1741 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat
     1801 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta
     1861 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc
     1921 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt
     1981 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg
     2041 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc
     2101 ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc
     2161 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct
     2221 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcg
     2281 tcgactttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa
     2341 aaggggggac tggaagggct aattcactcc caacgaagac aagatctgct ttttgcttgt
     2401 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
     2461 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
     2521 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
     2581 agcagtacgt atagtagttc atgtcatctt attattcagt atttataact tgcaaagaaa
     2641 tgaatatcag agagtgagag gaacttgttt attgcagctt ataatggtta caaataaagc
     2701 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     2761 tccaaactca tcaatgtatc ttatcatgtc tggctctagc tatcccgccc ctaactccgc
     2821 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     2881 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     2941 gaggcttttt tggaggccta gggacgtacc caattcgccc tatagtgagt cgtattacgc
     3001 gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact
     3061 taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac
     3121 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg
     3181 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     3241 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     3301 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     3361 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     3421 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     3481 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     3541 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     3601 aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     3661 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     3721 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     3781 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     3841 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     3901 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     3961 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     4021 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     4081 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     4141 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     4201 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     4261 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     4321 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     4381 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     4441 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     4501 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     4561 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     4621 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     4681 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     4741 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     4801 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     4861 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     4921 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     4981 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     5041 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     5101 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     5161 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     5221 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     5281 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     5341 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     5401 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     5461 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     5521 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
     5581 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
     5641 agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac
     5701 tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga
     5761 aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag
     5821 ctggagctgc aagcttaatg tagtcttatg caatactctt gtagtcttgc aacatggtaa
     5881 cgatgagtta gcaacatgcc ttacaaggag agaaaaagca ccgtgcatgc cgattggtgg
     5941 aagtaaggtg gtacgatcgt gccttattag gaaggcaaca gacgggtctg acatggattg
     6001 gacgaaccac tgaattgccg cattgcagag atattgtatt taagtgccta gctcgataca
     6061 taaacgggtc tctctggtta gaccagatct gagcctggga gctctctggc taactaggga
     6121 acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc
     6181 tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct
     6241 ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg agctctctcg
     6301 acgcaggact cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc gactggtgag
     6361 tacgccaaaa attttgacta gcggaggcta gaaggagaga gatgggtgcg agagcgtcag
     6421 tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc cagggggaaa
     6481 gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac gattcgcagt
     6541 taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac agctacaacc
     6601 atcccttcag acaggatcag aagaacttag atcattatat aatacagtag caaccctcta
     6661 ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca agatagagga
     6721 agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc gctgatcttc agacctggag
     6781 gaggagatat gagggacaat tggagaagtg aattatataa atataaagta gtaaaaattg
     6841 aaccattagg agtagcaccc accaaggcaa agagaagagt ggtgcagaga gaaaaaagag
     6901 cagtgggaat aggagctttg ttccttgggt tcttgggagc agcaggaagc actatgggcg
     6961 cagcgtcaat gacgctgacg gtacaggcca gacaattatt gtctggtata gtgcagcagc
     7021 agaacaattt gctgagggct attgaggcgc aacagcatct gttgcaactc acagtctggg
     7081 gcatcaagca gctccaggca agaatcctgg ctgtggaaag atacctaaag gatcaacagc
     7141 tcctggggat ttggggttgc tctggaaaac tcatttgcac cactgctgtg ccttggaatg
     7201 ctagttggag taataaatct ctggaacaga tttggaatca cacgacctgg atggagtggg
     7261 acagagaaat taacaattac acaagcttaa tacactcctt aattgaagaa tcgcaaaacc
     7321 agcaagaaaa gaatgaacaa gaattattgg aattagataa atgggcaagt ttgtggaatt
     7381 ggtttaacat aacaaattgg ctgtggtata taaaattatt cataatgata gtaggaggct
     7441 tggtaggttt aagaatagtt tttgctgtac tttctatagt gaatagagtt aggcagggat
     7501 attcaccatt atcgtttcag acccacctcc caaccccgag gggacccaga gagggcctat
     7561 ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag ataattagaa
     7621 ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga aagtaataat
     7681 ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat atgcttaccg
     7741 taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga cgaaacaccg
     7801 gagacgttga ctatcgtctc gctactctac cacttgtact tcagcggtca gcttactcga
     7861 cttaacgtgc acgtgacacg ttctagaccg tacatgctta catgggatga agcttggcgt
     7921 aactagatct tgagacaaat ggcagtattc atccacaatt ttaaaagaaa aggggggatt
     7981 ggggggtaca gtgcagggga aagaatagta gacataatag caacagacat acaaactaaa
     8041 gaattacaaa aacaaattac aaaaattcaa aattttcggg tttattacag ggacagcaga
     8101 gatccacttt ggcgccggct cgagggggc
//
