LOCUS       sequence-402892-        9150 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lineage tracing vector (with barcoded Target Sites and triple 
            sgRNAs).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9150)
  AUTHORS   Yang D, Jones MG, Naranjo S, Rideout WM 3rd, Min KHJ, Ho R, Wu W, 
            Replogle JM, Page JL, Quinn JJ, Horns F, Qiu X, Chen MZ, 
            Freed-Pastor WA, McGinnis CS, Patterson DM, Gartner ZJ, Chow ED, 
            Bivona TG, Chan MM, Yosef N, Jacks T, Weissman JS
  TITLE     Lineage tracing reveals the phylodynamics, plasticity, and paths of 
            tumor evolution.
  JOURNAL   Cell. 2022 Apr 28. pii: S0092-8674(22)00462-7. doi: 
            10.1016/j.cell.2022.04.015.
  PUBMED    35523183
REFERENCE   2  (bases 1 to 9150)
  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: "Cell. 2022 
            Apr 28. pii: S0092-8674(22)00462-7. doi: 
            10.1016/j.cell.2022.04.015."
COMMENT     Sequence Label: pDYT001
FEATURES             Location/Qualifiers
     source          1..9150
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(268..289)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     480..499
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     713..735
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     727..744
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   874..936
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        2065..3242
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2296..3234
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3191..3211
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             3374..4084
                     /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(3519..3537)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3787..3806
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    4412..4533
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4449..4468)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4503..4522
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5445..5465
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     5690..5710
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     5861..5880
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     repeat_region   6604..6638
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(7129..7145)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7129..7145)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7142..7164)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7153..7169
                     /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)."
     promoter        complement(join(7177..7183,7184..7201,7202..7207))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7222..7243
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7360..7377)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7531..8119)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7611..7630)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8290..9081,9082..9150))
                     /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     8913..8932
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgt gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaac aaaaactcaa
     1081 aatttcttct ataaagtaac aaaactttta tgagggacag ccccccccca aagcccccag
     1141 ggatgtaatt acgtccctcc cccgctaggg ggcagcagcg agccgcccgg ggctccgctc
     1201 cggtccggcg ctccccccgc atccccgagc cggcagcgtg cggggacagc ccgggcacgg
     1261 ggaaggtggc acgggatcgc tttcctctga acgcttctcg ctgctctttg agcctgcaga
     1321 cacctggggg gatacgggga aaaggcctcc acggccaagg atctgcgata tcgataagct
     1381 tgatatcgaa ttcgggaggt ggtccctgca gttacgccaa tgataacccc cgccagaaaa
     1441 atcttagtag ccttcccttt ttgttttccg tgccccaact cggcggattg actcggcccc
     1501 ttccggaaac acccgaatca acttctagtc aaattattgt tcacgccgca atgacccacc
     1561 cctggcccgc gtctgtggaa ctgacccctg gtgtacagga gagttcgctg ctgaaagtgg
     1621 tcccaaaggg gtactagttt ttaagctccc aactccccct cccccagcgt ctggaggatt
     1681 ccacaccctc gcaccgcagg ggcgaggaag tgggcggagt ccggttttgg cgccagccgc
     1741 tgaggctgcc aagcagaaaa gccaccgctg aggagactcc ggtcactgtc ctcgccccgc
     1801 ctcccccttc cctccccttg gggaccaccg ggcgccacgc cgcgaacggt aagtgccgcg
     1861 gtcgtcggcg cctccgccct ccccctaggg ccccaattcc cagcgggcgc ggcgcgcggc
     1921 ccctcccccc gccgggcgcg cgcccgctgc cccgcccttc gtggccgccc ggcgtgggcg
     1981 gtgccacccc tccccccggc ggccccgcgc gcagctcccg gctccctccc ccttcggatg
     2041 tggcttgagc tgtaggcgcg gaggggctcc ggtgcccgtc agtgggcaga gcgcacatcg
     2101 cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt gaaccggtgc ctagagaagg
     2161 tggcgcgggg taaactggga aagtgatgtc gtgtactggc tccgcctttt tcccgagggt
     2221 gggggagaac cgtatataag tgcagtagtc gccgtgaacg ttctttttcg caacgggttt
     2281 gccgccagaa cacaggtaag tgccgtgtgt ggttcccgcg ggcctggcct ctttacgggt
     2341 tatggccctt gcgtgccttg aattacttcc acctggctgc agtacgtgat tcttgatccc
     2401 gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt gcgcttaagg agccccttcg
     2461 cctcgtgctt gagttgaggc ctggcctggg cgctggggcc gccgcgtgcg aatctggtgg
     2521 caccttcgcg cctgtctcgc tgctttcgat aagtctctag ccatttaaaa tttttgatga
     2581 cctgctgcga cgcttttttt ctggcaagat agtcttgtaa atgcgggcca agatctgcac
     2641 actggtattt cggtttttgg ggccgcgggc ggcgacgggg cccgtgcgtc ccagcgcaca
     2701 tgttcggcga ggcggggcct gcgagcgcgg ccaccgagaa tcggacgggg gtagtctcaa
     2761 gctggccggc ctgctctggt gcctggcctc gcgccgccgt gtatcgcccc gccctgggcg
     2821 gcaaggctgg cccggtcggc accagttgcg tgagcggaaa gatggccgct tcccggccct
     2881 gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag agcgggcggg tgagtcaccc
     2941 acacaaagga aaagggcctt tccgtcctca gccgtcgctt catgtgactc cacggagtac
     3001 cgggcgccgt ccaggcacct cgattagttc tcgagctttt ggagtacgtc gtctttaggt
     3061 tggggggagg ggttttatgc gatggagttt ccccacactg agtgggtgga gactgaagtt
     3121 aggccagctt ggcacttgat gtaattctcc ttggaatttg ccctttttga gtttggatct
     3181 tggttcattc tcaagcctca gacagtggtt caaagttttt ttcttccatt tcaggtgtcg
     3241 tggaattctc acgcgtctgc agctagctat ggtgagcaag ggcgacactg ctcgatccgc
     3301 tcgcacctga ctgctacgtg actacgctga tgtacagatc catactgcat gttcattgac
     3361 gatctaccta gctatggtga gcaagggcga ggaggataac atggccatca tcaaggagtt
     3421 catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg
     3481 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
     3541 cggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc
     3601 ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt
     3661 caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc
     3721 ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc
     3781 cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta
     3841 ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg
     3901 ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg
     3961 cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt
     4021 ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa
     4081 gtaatgaatt aattaagtca cgaatccagc tagctgtgca gctaatggac ccggcaattc
     4141 aactgcagta atgctacctc gtactcacgc tttccaagtg cttggcgtcg catctcggtc
     4201 ctttgtacgc cgaaaaatgg cctgacaact aagctacggc acgctgccat gttgggtcat
     4261 aacgatatct ctggttcatc cgtgaccgaa catgtaatgg agtagcagga gctattaatt
     4321 cgcggaggac aatgcggttc gtagtcactg tcttccgcaa tcgtccatcg ctcctgcagg
     4381 tgttaactgc tcacctatta gcggctaagg aaacttgttt attgcagctt ataatggtta
     4441 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     4501 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggaattgac tcaaatgatg
     4561 tcaattagtc tatcagaagc tcatctggtc tcccttccgg gggacaagac atccctgttt
     4621 aatatttaaa cagcagtgtt cccaaactgg gttcttatat cccttgctct ggtcaaccag
     4681 gttgcagggt ttcctgtcct cacaggaacg aagtccctaa agaaacagtg gcagccaggt
     4741 ttagccccgg aattgactgg attccttttt tagggcccat tggtctagac gtgaccgagc
     4801 ttgtctgccc gcgcagtcca ctagacagac gcgcagaggt cgggcgggcc tagggccaat
     4861 ttcccatggt tcccttcatt tgcatatatg atgtaataag gttatggaga ctattaaact
     4921 tagccctaat caaactatat gatgataatg tgtgtggtac aaaaggtcat aacttattat
     4981 atactttgaa acttaaaaaa gggttacagt ttagtcacca taactgtaaa attttttcta
     5041 ttcttagctt tatatagttc ttgagaggcc atgtttatgg cagaagctat taattcgcgg
     5101 gtttaagagc taagctggaa acagcatagc aagtttaaat aaggctagtc cgttatcaac
     5161 ttgaaaaagt ggcaccgagt cggtgctttt tttccagtct gactatcacg atggatccga
     5221 cgcgccatct ctaggcccgc gccggccccc tcgcacggac ttgtgggaga agctcggcta
     5281 ctcccctgcc ccggttaatt tgcatataat atttcctagt aactatagag gcttaatgtg
     5341 cgataaaaga cagataatct gttcttttta atactagcta cattttacat gataggcttg
     5401 gatttctata agagatacaa atactaaatt attattttaa aaaacagcac aaaaggaaac
     5461 tcaccctaac tgtaaagtaa ttgtgtgttt tgagactata agtatccctt ggagaaccac
     5521 cttgttggtc ctttgtatgc cgaaaaagtt tgagagctaa gcagaaagct gcatagcaag
     5581 ttcaaataag gctagtccgt acacaacttg aaaaagtggc agccgagtcg gctgcttttt
     5641 ttgctcgaat ctacactcag ctatggcgcg ccccaaggtc gggcaggaag agggcctatt
     5701 tcccatgatt ccttcatatt tgcatatacg atacaaggct gttagagaga taattggaat
     5761 taatttgact gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt
     5821 tcttgggtag tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt
     5881 aacttgaaag tatttcgatt tcttggcttt atatatcttg tggaaagcca gaaacatggt
     5941 tggatcataa cgatatctcg tttcagagct aagcacaaga gtgcatagca agttgaaata
     6001 aggctagtcc gtttacaact tgaaaaagtg gcacccgagt cgggtgcttt ttttctcgag
     6061 tactaggatc cattaggcgg ccgcgtttaa acatggcttt ttccccgtat ccccccaggt
     6121 gtctgcaggc tcaaagagca gcgagaagcg ttcagaggaa agcgatcccg tgccaccttc
     6181 cccgtgcccg ggctgtcccc gcacgctgcc ggctcgggga tgcgggggga gcgccggacc
     6241 ggagcggagc cccgggcggc tcgctgctgc cccctagcgg gggagggacg taattacatc
     6301 cctgggggct ttgggggggg gctgtccctg atatctataa caagaaaata tatatataat
     6361 aagttatcac gtaagtagaa catgaaataa caatataatt atcgtatgag ttaaatctta
     6421 aaagtcacgt aaaagataat catgcgtcat tttgactcac gcggtcgtta tagttcaaaa
     6481 tcagtgacac ttaccgcatt gacaagcacg cctcacggga gctccaagcg gcgactgaga
     6541 tgtcctaaat gcacagcgac ggattcgcgc tatttagaaa gagagagcaa tatttcaaga
     6601 atgcatgcgt caattttacg cagactatct ttctagggtt aatctagctg catcaggatc
     6661 atatcgtcgg gtcttttttc cggctcagtc atcgcccaag ctggcgctat ctgggcatcg
     6721 gggaggaaga agcccgtgcc ttttcccgcg aggttgaagc ggcatggaaa gagtttgccg
     6781 aggatgactg ctgctgcatt gacgttgagc gaaaacgcac gtttaccatg atgattcggg
     6841 aaggtgtggc catgcacgcc tttaacggtg aactgttcgt tcaggccacc tgggatacca
     6901 gttcgtcgcg gcttttccgg acacagttcc ggatggtcag cccgaagcgc atcagcaacc
     6961 cgaacaatac cggcgacagc cggaactgcc gtgccggtgt gcagattaat gacagcggtg
     7021 cggcgctggg atattacgtc agcgaggacg ggtatcctgg ctggatgccg cagaaatgga
     7081 catggatacc ccgtgagtta cccggcgggc gcgcttggcg taatcatggt catagctgtt
     7141 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     7201 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     7261 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     7321 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     7381 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     7441 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     7501 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     7561 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     7621 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     7681 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     7741 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     7801 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     7861 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     7921 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     7981 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     8041 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     8101 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     8161 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     8221 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     8281 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     8341 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     8401 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     8461 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     8521 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     8581 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     8641 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     8701 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     8761 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     8821 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     8881 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     8941 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     9001 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     9061 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     9121 aagggcgaca cggaaatgtt gaatactcat
//
