LOCUS       sequence-207222-        8309 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector that contains an optimized S. pyogenes sgRNA 
            targeting human telomeres.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8309)
  AUTHORS   Chen B, Gilbert LA, Cimini BA, Schnitzbauer J, Zhang W, Li GW, Park 
            J, Blackburn EH, Weissman JS, Qi LS, Huang B
  TITLE     Dynamic Imaging of Genomic Loci in Living Human Cells by an 
            Optimized CRISPR/Cas System.
  JOURNAL   Cell. 2013 Dec 19;155(7):1479-91. doi: 10.1016/j.cell.2013.12.001.
  PUBMED    24360272
REFERENCE   2  (bases 1 to 8309)
  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"; doi: 
            "10.1016/j.cell.2013.12"; journalName: "Cell"; date: "2013-12-19- 
            19"; volume: "155"; issue: "7"; pages: "1479-91"
COMMENT     Sequence Label: pSLQ1651-sgTelomere(F+E)
FEATURES             Location/Qualifiers
     source          1..8309
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(40..59)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        231..610
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        612..810
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             828..1008
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1055..1180
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1677..1910
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2095..2139
                     /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    2437..2554
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    complement(2807..2840)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     2844..2864
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     enhancer        3159..3462
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3463..3666
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             3700..4296
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3700..3719)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4196..4216
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             4306..4359
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     regulatory      4366..4375
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4372..5082
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4517..4535)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4785..4804
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(5102..5135)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    5191..5779
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5244..5264)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5662..5673)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5782..5798)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5783..5799)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             6308..6488
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6550..7138)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6630..6649)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7309..8100,8101..8169))
                     /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     7932..7951
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8170..8274)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gatcgggaga tctcccgatc ccctatggtg cactctcagt acaatctgct ctgatgccgc
       61 atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag
      121 caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag
      181 ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt gacattgatt
      241 attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga
      301 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
      361 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
      421 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
      481 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
      541 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
      601 tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
      661 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
      721 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
      781 gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg tctctctggt
      841 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
      901 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
      961 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa
     1021 cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga ctcggcttgc
     1081 tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa aaattttgac
     1141 tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc gggggagaat
     1201 tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat ataaattaaa
     1261 acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg gcctgttaga
     1321 aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc agacaggatc
     1381 agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc atcaaaggat
     1441 agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa acaaaagtaa
     1501 gaccaccgca cagcaagcgg ccggccgctg atcttcagac ctggaggagg agatatgagg
     1561 gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc attaggagta
     1621 gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt gggaatagga
     1681 gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc gtcaatgacg
     1741 ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa caatttgctg
     1801 agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat caagcagctc
     1861 caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct ggggatttgg
     1921 ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag ttggagtaat
     1981 aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag agaaattaac
     2041 aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca agaaaagaat
     2101 gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt taacataaca
     2161 aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt aggtttaaga
     2221 atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc accattatcg
     2281 tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat agaagaagaa
     2341 ggtggagaga gagacagaga cagatccatt cgattagtga acggatcggc actgcgtgcg
     2401 ccaattctgc agacaaatgg cagtattcat ccacaatttt aaaagaaaag gggggattgg
     2461 ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac aaactaaaga
     2521 attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg acagcagaga
     2581 tccagtttgg ttagtaccgg gcccgctcta gagatccgac gcgccatctc taggcccgcg
     2641 ccggccccct cgcacggact tgtgggagaa gctcggctac tcccctgccc cggttaattt
     2701 gcatataata tttcctagta actatagagg cttaatgtgc gataaaagac agataatctg
     2761 ttctttttaa tactagctac attttacatg ataggcttgg atttctataa cttcgtatag
     2821 catacattat acgaagttat aaacagcaca aaaggaaact caccctaact gtaaagtaat
     2881 tgtgtgtttt gagactataa gtatcccttg gagaaccacc ttgttggtta gggttagggt
     2941 tagggttagt ttaagagcta tgctggaaac agcatagcaa gtttaaataa ggctagtccg
     3001 ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt tctcgagtac taggatccat
     3061 taggcggccg cgtggataac cgtattaccg ccatgcatta gttattaata gtaatcaatt
     3121 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     3181 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     3241 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     3301 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     3361 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     3421 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     3481 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     3541 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     3601 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     3661 agagctggtt tagtgaaccg tcagatccgc tagcccacca tgaccgagta caagcccacg
     3721 gtgcgcctcg ccacccgcga cgacgtcccc cgggccgtac gcaccctcgc cgccgcgttc
     3781 gccgactacc ccgccacgcg ccacaccgtc gacccggacc gccacatcga gcgggtcacc
     3841 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     3901 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     3961 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     4021 cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     4081 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     4141 gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac
     4201 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     4261 ccgcgcacct ggtgcatgac ccgcaagccc ggtgccggat cgggagaggg cagaggaagt
     4321 ctgctaacat gcggtgacgt cgaggagaat cctggcccac cggtcgccac catggtgagc
     4381 aagggcgagg aggataacat ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg
     4441 gagggctccg tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac
     4501 gagggcaccc agaccgccaa gctgaaggtg accaagggtg gccccctgcc cttcgcctgg
     4561 gacatcctgt cccctcagtt catgtacggc tccaaggcct acgtgaagca ccccgccgac
     4621 atccccgact acttgaagct gtccttcccc gagggcttca agtgggagcg cgtgatgaac
     4681 ttcgaggacg gcggcgtggt gaccgtgacc caggactcct ccctgcagga cggcgagttc
     4741 atctacaagg tgaagctgcg cggcaccaac ttcccctccg acggccccgt aatgcagaag
     4801 aagaccatgg gctgggaggc ctcctccgag cggatgtacc ccgaggacgg cgccctgaag
     4861 ggcgagatca agcagaggct gaagctgaag gacggcggcc actacgacgc tgaggtcaag
     4921 accacctaca aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt caacatcaag
     4981 ttggacatca cctcccacaa cgaggactac accatcgtgg aacagtacga acgcgccgag
     5041 ggccgccact ccaccggcgg catggacgag ctgtacaagt aagaattcgt cgagggacct
     5101 aataacttcg tatagcatac attatacgaa gttatacatg tttaagggtt ccggttccac
     5161 taggtacaat tcgatatcaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa
     5221 agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta
     5281 atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa
     5341 tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg
     5401 tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc
     5461 ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc
     5521 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg
     5581 gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg
     5641 acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg
     5701 ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc
     5761 ctttgggccg cctccccgca tcgataccgt cgacctcgat cgagacctag aaaaacatgg
     5821 agcaatcaca agtagcaata cagcagctac caatgctgat tgtgcctggc tagaagcaca
     5881 agaggaggag gaggtgggtt ttccagtcac acctcaggta cctttaagac caatgactta
     5941 caaggcagct gtagatctta gccacttttt aaaagaaaag gggggactgg aagggctaat
     6001 tcactcccaa cgaagacaag atatccttga tctgtggatc taccacacac aaggctactt
     6061 ccctgattgg cagaactaca caccagggcc agggatcaga tatccactga cctttggatg
     6121 gtgctacaag ctagtaccag ttgagcaaga gaaggtagaa gaagccaatg aaggagagaa
     6181 cacccgcttg ttacaccctg tgagcctgca tgggatggat gacccggaga gagaagtatt
     6241 agagtggagg tttgacagcc gcctagcatt tcatcacatg gcccgagagc tgcatccgga
     6301 ctgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg
     6361 gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg
     6421 tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat
     6481 ctctagcagc atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     6541 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     6601 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     6661 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     6721 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     6781 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     6841 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     6901 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     6961 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     7021 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     7081 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     7141 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     7201 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     7261 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     7321 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     7381 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     7441 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     7501 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     7561 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     7621 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     7681 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     7741 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     7801 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     7861 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     7921 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     7981 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     8041 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     8101 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     8161 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     8221 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     8281 tccccgaaaa gtgccacctg acgtcgacg
//
