LOCUS       sequence-345343-        6268 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes 5xTAPE-1 construct that can be integrated using piggyBAC 
            system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6268)
  AUTHORS   Choi J, Chen W, Minkina A, Chardon FM, Suiter CC, Regalado SG, 
            Domcke S, Hamazaki N, Lee C, Martin B, Daza RM, Shendure J
  TITLE     A time-resolved, multi-symbol molecular recorder via sequential 
            genome editing.
  JOURNAL   Nature. 2022 Jul 6. pii: 10.1038/s41586-022-04922-8. doi: 
            10.1038/s41586-022-04922-8.
  PUBMED    35794474
REFERENCE   2  (bases 1 to 6268)
  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: "Nature. 
            2022 Jul 6. pii: 10.1038/s41586-022-04922-8. doi: 
            10.1038/s41586-022-04922-8."
COMMENT     Sequence Label: piggyBAC-5xTAPE-1-BlastR
FEATURES             Location/Qualifiers
     source          1..6268
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             164..432
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     CDS             458..853
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     primer_bind     complement(913..933)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    975..1096
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1012..1031)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1066..1085
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   1859..1893
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(2384..2400)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2384..2400)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2397..2419)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2408..2424
                     /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(2432..2438,2439..2456,2457..2462))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2477..2498
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2615..2632)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2786..3374)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2866..2885)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3545..4336,4337..4405))
                     /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     4168..4187
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4406..4510)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4536..4991)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4673..4694)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4885..4904
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5118..5140
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5132..5149
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5133..5149
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   5279..5341
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        5766..6014
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     5766..5786
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     5937..5956
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        join(6208..6268,1..151)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
ORIGIN
        1 gggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg
       61 cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct
      121 ttttcgcaac gggtttgccg ccagaacaca gctgaagctt cgaggggctc gcatctctcc
      181 ttcacgcgcc cgccgcccta cctgaggccg ccatccacgc cggttgagtc gcgttctgcc
      241 gcctcccgcc tgtggtgcct cctgaactgc gtccgccgtc taggtaagtt taaagctcag
      301 gtcgagaccg ggcctttgtc cggcgctccc ttggagccta cctagactca gccggctctc
      361 cacgctttgc ctgaccctgc ttgctcaact ctacgtcttt gtttcgtttt ctgttctgcg
      421 ccgttacaga tccaagctgt gaccggcgcc taccgtcatg gccaagcctt tgtctcaaga
      481 agaatccacc ctcattgaaa gagcaacggc tacaatcaac agcatcccca tctctgaaga
      541 ctacagcgtc gccagcgcag ctctctctag cgacggccgc atcttcactg gtgtcaatgt
      601 atatcatttt actgggggac cttgtgcaga actcgtggtg ctgggcactg ctgctgcggc
      661 agctggcaac ctgacttgta tcgtcgcgat cggaaatgag aacaggggca tcttgagccc
      721 ctgcggacgg tgccgacagg tgcttctcga tctgcatcct gggatcaaag ccatagtgaa
      781 ggacagtgat ggacagccga cggcagttgg gattcgtgaa ttgctgccct ctggttatgt
      841 gtgggagggc taaacgcgta atcaacctct ggattacaaa atttgtgaaa gattgactgg
      901 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
      961 tcatgcgtta actaaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     1021 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     1081 tcatcaatgt atcttatcat gtctggaatt gactcaaatg atgtcaatta gtctatcaga
     1141 agctcatctg gtctcccttc cgggggacaa gacatccctg tttaatattt aaacagcagt
     1201 gttcccaaac tgggttctta tatcccttgc tctggtcaac caggttgcag ggtttcctgt
     1261 cctcacagga acgaagtccc taaagaaaca gtggcagcca ggtttagccc cggaattgac
     1321 tggattcctt ttttagggcc cattggtatg gctttttccc cgtatccccc caggtgtctg
     1381 caggctcaaa gagcagcgag aagcgttcag aggaaagcga tcccgtgcca ccttccccgt
     1441 gcccgggctg tccccgcacg ctgccggctc ggggatgcgg ggggagcgcc ggaccggagc
     1501 ggagccccgg gcggctcgct gctgccccct agcgggggag ggacgtaatt acatccctgg
     1561 gggctttggg ggggggctgt ccctgatatc tataacaaga aaatatatat ataataagtt
     1621 atcacgtaag tagaacatga aataacaata taattatcgt atgagttaaa tcttaaaagt
     1681 cacgtaaaag ataatcatgc gtcattttga ctcacgcggt cgttatagtt caaaatcagt
     1741 gacacttacc gcattgacaa gcacgcctca cgggagctcc aagcggcgac tgagatgtcc
     1801 taaatgcaca gcgacggatt cgcgctattt agaaagagag agcaatattt caagaatgca
     1861 tgcgtcaatt ttacgcagac tatctttcta gggttaatct agctgcatca ggatcatatc
     1921 gtcgggtctt ttttccggct cagtcatcgc ccaagctggc gctatctggg catcggggag
     1981 gaagaagccc gtgccttttc ccgcgaggtt gaagcggcat ggaaagagtt tgccgaggat
     2041 gactgctgct gcattgacgt tgagcgaaaa cgcacgttta ccatgatgat tcgggaaggt
     2101 gtggccatgc acgcctttaa cggtgaactg ttcgttcagg ccacctggga taccagttcg
     2161 tcgcggcttt tccggacaca gttccggatg gtcagcccga agcgcatcag caacccgaac
     2221 aataccggcg acagccggaa ctgccgtgcc ggtgtgcaga ttaatgacag cggtgcggcg
     2281 ctgggatatt acgtcagcga ggacgggtat cctggctgga tgccgcagaa atggacatgg
     2341 ataccccgtg agttacccgg cgggcgcgct tggcgtaatc atggtcatag ctgtttcctg
     2401 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     2461 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     2521 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     2581 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     2641 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     2701 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     2761 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     2821 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     2881 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     2941 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3001 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3061 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3121 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3181 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     3241 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3301 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3361 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3421 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     3481 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     3541 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     3601 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     3661 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     3721 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     3781 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     3841 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     3901 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     3961 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     4021 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     4081 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     4141 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     4201 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     4261 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     4321 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     4381 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     4441 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     4501 gggttccgcg cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt
     4561 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     4621 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     4681 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     4741 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     4801 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     4861 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     4921 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     4981 acagggcgcg tcccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     5041 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     5101 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg agcgcgcctc
     5161 gttcattcac gtttttgaac ccgtggagga cgggcagact cgcggtgcaa atgtgtttta
     5221 cagcgtgatg gagcagatga agatgctcga cacgctgcag aacacgcagc tagattaacc
     5281 ctagaaagat aatcatattg tgacgtacgt taaagataat catgtgtaaa attgacgcat
     5341 gtgttttatc ggtctgtata tcgaggttta tttattaatt tgaatagata ttaagtttta
     5401 ttatatttac acttacatac taataataaa ttcaacaaac aatttattta tgtttattta
     5461 tttattaaaa aaaacaaaaa ctcaaaattt cttctataaa gtaacaaaac ttttatgagg
     5521 gacagccccc ccccaaagcc cccagggatg taattacgtc cctcccccgc tagggggcag
     5581 cagcgagccg cccggggctc cgctccggtc cggcgctccc cccgcatccc cgagccggca
     5641 gcgtgcgggg acagcccggg cacggggaag gtggcacggg atcgctttcc tctgaacgct
     5701 tctcgctgct ctttgagcct gcagacacct ggggggatac ggggaaaagg cctccacggc
     5761 caagagaggg cctatttccc atgattcctt catatttgca tatacgatac aaggctgtta
     5821 gagagataat tagaattaat ttgactgtaa acacaaagat attagtacaa aatacgtgac
     5881 gtagaaagta ataatttctt gggtagtttg cagttttaaa attatgtttt aaaatggact
     5941 atcatatgct taccgtaact tgaaagtatt tcgatttctt ggctttatat atcttgtgga
     6001 aaggacgaaa caccttacct atggatgatg gtgagcacgt gatggtgagc acgtgatggt
     6061 gagcacgtga tggtgagcac gtgatggtga gcacgtgatg gtgagcgctt taaggccggt
     6121 cctagcaatt tttttaagct tggcgtaact agatcttgag acaaacgcgg ccgcgaagga
     6181 tctgcgatcg ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga
     6241 agttgggggg aggggtcggc aattgaac
//
