LOCUS       sequence-402885-        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: pDYT002
FEATURES             Location/Qualifiers
     source          1..9150
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     208..227
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(446..550)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(576..1031)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(713..734)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     925..944
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1158..1180
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1172..1189
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1173..1189
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   1319..1381
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        2510..3687
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2741..3679
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3636..3656
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             3819..4529
                     /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(3964..3982)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4232..4251
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    4857..4978
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4894..4913)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4948..4967
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5890..5910
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     6135..6155
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6306..6325
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     repeat_region   7049..7083
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(7574..7590)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7574..7590)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7587..7609)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7598..7614
                     /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(7622..7628,7629..7646,7647..7652))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7667..7688
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7805..7822)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7976..8564)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8056..8075)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8735..9150,1..376,377..445))
                     /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"
ORIGIN
        1 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
       61 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
      121 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
      181 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
      241 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
      301 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
      361 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
      421 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
      481 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
      541 gggttccgcg cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt
      601 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
      661 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
      721 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
      781 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
      841 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
      901 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
      961 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     1021 acagggcgcg tcccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     1081 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     1141 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg agcgcgcctc
     1201 gttcattcac gtttttgaac ccgtggagga cgggcagact cgcggtgcaa atgtgtttta
     1261 cagcgtgatg gagcagatga agatgctcga cacgctgcag aacacgcagc tagattaacc
     1321 ctagaaagat aatcatattg tgacgtacgt taaagataat catgtgtaaa attgacgcat
     1381 gtgttttatc ggtctgtata tcgaggttta tttattaatt tgaatagata ttaagtttta
     1441 ttatatttac acttacatac taataataaa ttcaacaaac aatttattta tgtttattta
     1501 tttattaaaa aaaacaaaaa ctcaaaattt cttctataaa gtaacaaaac ttttatgagg
     1561 gacagccccc ccccaaagcc cccagggatg taattacgtc cctcccccgc tagggggcag
     1621 cagcgagccg cccggggctc cgctccggtc cggcgctccc cccgcatccc cgagccggca
     1681 gcgtgcgggg acagcccggg cacggggaag gtggcacggg atcgctttcc tctgaacgct
     1741 tctcgctgct ctttgagcct gcagacacct ggggggatac ggggaaaagg cctccacggc
     1801 caaggatctg cgatatcgat aagcttgata tcgaattcgg gaggtggtcc ctgcagttac
     1861 gccaatgata acccccgcca gaaaaatctt agtagccttc cctttttgtt ttccgtgccc
     1921 caactcggcg gattgactcg gccccttccg gaaacacccg aatcaacttc tagtcaaatt
     1981 attgttcacg ccgcaatgac ccacccctgg cccgcgtctg tggaactgac ccctggtgta
     2041 caggagagtt cgctgctgaa agtggtccca aaggggtact agtttttaag ctcccaactc
     2101 cccctccccc agcgtctgga ggattccaca ccctcgcacc gcaggggcga ggaagtgggc
     2161 ggagtccggt tttggcgcca gccgctgagg ctgccaagca gaaaagccac cgctgaggag
     2221 actccggtca ctgtcctcgc cccgcctccc ccttccctcc ccttggggac caccgggcgc
     2281 cacgccgcga acggtaagtg ccgcggtcgt cggcgcctcc gccctccccc tagggcccca
     2341 attcccagcg ggcgcggcgc gcggcccctc cccccgccgg gcgcgcgccc gctgccccgc
     2401 ccttcgtggc cgcccggcgt gggcggtgcc acccctcccc ccggcggccc cgcgcgcagc
     2461 tcccggctcc ctcccccttc ggatgtggct tgagctgtag gcgcggaggg gctccggtgc
     2521 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
     2581 caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
     2641 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
     2701 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc
     2761 ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg
     2821 gctgcagtac gtgattcttg atcccgagct tcgggttgga agtgggtggg agagttcgag
     2881 gccttgcgct taaggagccc cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg
     2941 gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc
     3001 tctagccatt taaaattttt gatgacctgc tgcgacgctt tttttctggc aagatagtct
     3061 tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt tttggggccg cgggcggcga
     3121 cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc
     3181 gagaatcgga cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc
     3241 gccgtgtatc gccccgccct gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc
     3301 ggaaagatgg ccgcttcccg gccctgctgc agggagctca aaatggagga cgcggcgctc
     3361 gggagagcgg gcgggtgagt cacccacaca aaggaaaagg gcctttccgt cctcagccgt
     3421 cgcttcatgt gactccacgg agtaccgggc gccgtccagg cacctcgatt agttctcgag
     3481 cttttggagt acgtcgtctt taggttgggg ggaggggttt tatgcgatgg agtttcccca
     3541 cactgagtgg gtggagactg aagttaggcc agcttggcac ttgatgtaat tctccttgga
     3601 atttgccctt tttgagtttg gatcttggtt cattctcaag cctcagacag tggttcaaag
     3661 tttttttctt ccatttcagg tgtcgtggaa ttctcacgcg tctgcagcta gctatggtga
     3721 gcaagggcga cactgctcga tccgctcgca cctgactgct acgtgactac gctgatgtac
     3781 agatccatac tgcatgttca ttgacgatct acctagctat ggtgagcaag ggcgaggagg
     3841 ataacatggc catcatcaag gagttcatgc gcttcaaggt gcacatggag ggctccgtga
     3901 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
     3961 ccgccaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac atcctgtccc
     4021 ctcagttcat gtacggctcc aaggcctacg tgaagcaccc cgccgacatc cccgactact
     4081 tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     4141 gcgtggtgac cgtgacccag gactcctccc tgcaggacgg cgagttcatc tacaaggtga
     4201 agctgcgcgg caccaacttc ccctccgacg gccccgtaat gcagaagaag accatgggct
     4261 gggaggcctc ctccgagcgg atgtaccccg aggacggcgc cctgaagggc gagatcaagc
     4321 agaggctgaa gctgaaggac ggcggccact acgacgctga ggtcaagacc acctacaagg
     4381 ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa catcaagttg gacatcacct
     4441 cccacaacga ggactacacc atcgtggaac agtacgaacg cgccgagggc cgccactcca
     4501 ccggcggcat ggacgagctg tacaagtaat gaattaatta agtcacgaat ccagctagct
     4561 gtgcagcagg cttagcgtac aattcaactg cagtaatgct acctcgtact cacgctttcc
     4621 aagtgcttgg cgtcgcatct cggtcctttg tacgccgaaa aatggcctga caactaagct
     4681 acggcacgct gccatgttgg gtcataacga tatctctggt tcatccgtga ccgaacatgt
     4741 catggagtag caggagctat taattcgcgg aggacaatgc ggttcgtagt cactgtcttc
     4801 cgcaatcgtc catcgctcct gcaggtgtta actgctcacc tattagcggc taaggaaact
     4861 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     4921 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     4981 atgtctggaa ttgactcaaa tgatgtcaat tagtctatca gaagctcatc tggtctccct
     5041 tccgggggac aagacatccc tgtttaatat ttaaacagca gtgttcccaa actgggttct
     5101 tatatccctt gctctggtca accaggttgc agggtttcct gtcctcacag gaacgaagtc
     5161 cctaaagaaa cagtggcagc caggtttagc cccggaattg actggattcc ttttttaggg
     5221 cccattggtc tagacgtgac cgagcttgtc tgcccgcgca gtccactaga cagacgcgca
     5281 gaggtcgggc gggcctaggg ccaatttccc atggttccct tcatttgcat atatgatgta
     5341 ataaggttat ggagactatt aaacttagcc ctaatcaaac tatatgatga taatgtgtgt
     5401 ggtacaaaag gtcataactt attatatact ttgaaactta aaaaagggtt acagtttagt
     5461 caccataact gtaaaatttt ttctattctt agctttatat agttcttgag aggccatgtt
     5521 tatggcagga gctatcaatt cgcgggttta agagctaagc tggaaacagc atagcaagtt
     5581 taaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg ctttttttcc
     5641 agtctgacta tcacgatgga tccgacgcgc catctctagg cccgcgccgg ccccctcgca
     5701 cggacttgtg ggagaagctc ggctactccc ctgccccggt taatttgcat ataatatttc
     5761 ctagtaacta tagaggctta atgtgcgata aaagacagat aatctgttct ttttaatact
     5821 agctacattt tacatgatag gcttggattt ctataagaga tacaaatact aaattattat
     5881 tttaaaaaac agcacaaaag gaaactcacc ctaactgtaa agtaattgtg tgttttgaga
     5941 ctataagtat cccttggaga accaccttgt tggtcctttg tatgccgaaa aagtttgaga
     6001 gctaagcaga aagctgcata gcaagttcaa ataaggctag tccgtacaca acttgaaaaa
     6061 gtggcagccg agtcggctgc tttttttgct cgaatctaca ctcagctatg gcgcgcccca
     6121 aggtcgggca ggaagagggc ctatttccca tgattccttc atatttgcat atacgataca
     6181 aggctgttag agagataatt ggaattaatt tgactgtaaa cacaaagata ttagtacaaa
     6241 atacgtgacg tagaaagtaa taatttcttg ggtagtttgc agttttaaaa ttatgtttta
     6301 aaatggacta tcatatgctt accgtaactt gaaagtattt cgatttcttg gctttatata
     6361 tcttgtggaa agccagaaac atggttgggt cataacgata cctcgtttca gagctaagca
     6421 caagagtgca tagcaagttg aaataaggct agtccgttta caacttgaaa aagtggcacc
     6481 cgagtcgggt gctttttttc tcgagtacta ggatccatta ggcggccgcg tttaaacatg
     6541 gctttttccc cgtatccccc caggtgtctg caggctcaaa gagcagcgag aagcgttcag
     6601 aggaaagcga tcccgtgcca ccttccccgt gcccgggctg tccccgcacg ctgccggctc
     6661 ggggatgcgg ggggagcgcc ggaccggagc ggagccccgg gcggctcgct gctgccccct
     6721 agcgggggag ggacgtaatt acatccctgg gggctttggg ggggggctgt ccctgatatc
     6781 tataacaaga aaatatatat ataataagtt atcacgtaag tagaacatga aataacaata
     6841 taattatcgt atgagttaaa tcttaaaagt cacgtaaaag ataatcatgc gtcattttga
     6901 ctcacgcggt cgttatagtt caaaatcagt gacacttacc gcattgacaa gcacgcctca
     6961 cgggagctcc aagcggcgac tgagatgtcc taaatgcaca gcgacggatt cgcgctattt
     7021 agaaagagag agcaatattt caagaatgca tgcgtcaatt ttacgcagac tatctttcta
     7081 gggttaatct agctgcatca ggatcatatc gtcgggtctt ttttccggct cagtcatcgc
     7141 ccaagctggc gctatctggg catcggggag gaagaagccc gtgccttttc ccgcgaggtt
     7201 gaagcggcat ggaaagagtt tgccgaggat gactgctgct gcattgacgt tgagcgaaaa
     7261 cgcacgttta ccatgatgat tcgggaaggt gtggccatgc acgcctttaa cggtgaactg
     7321 ttcgttcagg ccacctggga taccagttcg tcgcggcttt tccggacaca gttccggatg
     7381 gtcagcccga agcgcatcag caacccgaac aataccggcg acagccggaa ctgccgtgcc
     7441 ggtgtgcaga ttaatgacag cggtgcggcg ctgggatatt acgtcagcga ggacgggtat
     7501 cctggctgga tgccgcagaa atggacatgg ataccccgtg agttacccgg cgggcgcgct
     7561 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     7621 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     7681 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     7741 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     7801 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     7861 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     7921 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     7981 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     8041 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     8101 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     8161 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     8221 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     8281 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     8341 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     8401 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     8461 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     8521 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     8581 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     8641 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     8701 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     8761 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     8821 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     8881 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     8941 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     9001 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     9061 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     9121 gagttacatg atcccccatg ttgtgcaaaa
//
