LOCUS       sequence-450391-        5420 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty MTK PB cargo backbone marked with puromycin resistance..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5420)
  AUTHORS   Loveless BT, Carlson CK, Dentzel Helmy CA, Hu VJ, Ross SK, Demelo 
            MC, Murtaza A, Liang G, Ficht M, Singhai A, Pajoh-Casco MJ, Liu CC
  TITLE     Open-ended molecular recording of sequential cellular events into 
            DNA
  JOURNAL   bioRxiv 2021.11.05.467507
REFERENCE   2  (bases 1 to 5420)
  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: "bioRxiv 
            2021.11.05.467507"
COMMENT     Sequence Label: pTBL2368 MTK PB puro backbone
FEATURES             Location/Qualifiers
     source          1..5420
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             513..524
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             531..1244
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      1256..1327
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1343..1370
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    1806..1923
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1992..2203
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             2229..2828
                     /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(2229..2248)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2725..2745
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    2988..3109
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3025..3044)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3079..3098
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   3304..3366
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        complement(3417..3435)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             complement(3475..4290)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(3475..4290)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4198..4217
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4282..4301
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4515..5103
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5004..5023
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5215..5234
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5215..5233
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   5263..5297
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
ORIGIN
        1 ttttgaacta taacgaccgc gtgagtcaaa atgacgcatg attatctttt acgtgacttt
       61 taagatttaa ctcatacgat aattatattg ttatttcatg ttctacttac gtgataactt
      121 attatatata tattttcttg ttatagatat caactagaat gctagcatgg gcccagatct
      181 ccatgggccc tgaattcgca tctagactga tgagacgtgg tagagccaca aacagccggt
      241 acaagcaacg atctccagga ccatctgaat catgcgcgga tgacacgaac tcacgacggc
      301 gatcacagac attaacccac agtacagaca ctgcgacaac gtggcaattc gtcgcaatac
      361 aacggaaagt gaaacgtgat ttcatgcgtc attttgaaca ttttgtaaat cttatttaat
      421 aatgtgtgcg gcaattcaca tttaatttat gaatgttttc ttaacatcgc ggcaactcaa
      481 gaaacggcag gttcggatct tagctactag agaaagagga gaaatactag atgcgtaaag
      541 gcgaagagct gttcactggt gtcgtcccta ttctggtgga actggatggt gatgtcaacg
      601 gtcataagtt ttccgtgcgt ggcgagggtg aaggtgacgc aactaatggt aaactgacgc
      661 tgaagttcat ctgtactact ggtaaactgc cggttccttg gccgactctg gtaacgacgc
      721 tgacttatgg tgttcagtgc tttgctcgtt atccggacca tatgaagcag catgacttct
      781 tcaagtccgc catgccggaa ggctatgtgc aggaacgcac gatttccttt aaggatgacg
      841 gcacgtacaa aacgcgtgcg gaagtgaaat ttgaaggcga taccctggta aaccgcattg
      901 agctgaaagg cattgacttt aaagaggacg gcaatatcct gggccataag ctggaataca
      961 attttaacag ccacaatgtt tacatcaccg ccgataaaca aaaaaatggc attaaagcga
     1021 attttaaaat tcgccacaac gtggaggatg gcagcgtgca gctggctgat cactaccagc
     1081 aaaacactcc aatcggtgat ggtcctgttc tgctgccaga caatcactat ctgagcacgc
     1141 aaagcgttct gtctaaagat ccgaacgaga aacgcgatca tatggttctg ctggagttcg
     1201 taaccgcagc gggcatcacg catggtatgg atgaactgta caaatgacca ggcatcaaat
     1261 aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa
     1321 cgctctctac tagagtcaca ctggctcacc ttcgggtggg cctttctgcg tttatagctg
     1381 gaaatctgct cgtcagtggt gctcacactg acgaatcatg tacagatcat accgatgact
     1441 gcctggcgac tcacaactaa gcaagacagc cggaaccagc gccggcgaac accactgcat
     1501 atatggcata tcacaacagt ccacgtctca agcagttaca gagatgttac gaaccctgaa
     1561 caggttgact attaattgtg tctgcttgat gtggacacca ggtggcgctg gacatcagat
     1621 ttggagaggc agttgtctag ggaaccgggc tctgtgccag cgcaggaggc aggctggctc
     1681 tcctattcca gggatgctca tccaggaagg aaaggttgca tgctggacac actaacctta
     1741 cagcggtggg ctctatgggg tacctacgta tagatcttga gacaaatggc agtattcatc
     1801 cacaatttta aaagaaaagg ggggattggg gggtacagtg caggggaaag aatagtagac
     1861 ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa aattcaaaat
     1921 tttcgggttt attacaggga cagcagagat ccactttggc gccggctcga gtggctccgg
     1981 tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt
     2041 cggcaattga accggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt
     2101 gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc
     2161 cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca caggtgtcgt gacgcgggat
     2221 ccgccaccat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtccccc
     2281 gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg
     2341 acccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg
     2401 ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca
     2461 cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt
     2521 tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc
     2581 ccaaggagcc cgcgtggttc ctggccaccg tcggagtctc gcccgaccac cagggcaagg
     2641 gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg
     2701 ccttcctgga aacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg
     2761 tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg
     2821 gtgcctgact cgagatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     2881 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     2941 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     3001 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     3061 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     3121 taactagtta aaagttttgt tactttatag aagaaatttt gagtttttgt ttttttttaa
     3181 taaataaata aacataaata aattgtttgt tgaatttatt attagtatgt aagtgtaaat
     3241 ataataaaac ttaatatcta ttcaaattaa taaataaacc tcgatataca gaccgataaa
     3301 acacatgcgt caattttacg catgattatc tttaacgtac gtcacaatat gattatcttt
     3361 ctagggttaa tctagtatac gcgtatgcgg ccgcttaatt aatccagctt ttgttccctt
     3421 tagtgagggt taattccgag cggccgcgtg ttacaaccaa ttaaccaatt ctgattagaa
     3481 aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat caataccata
     3541 tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt tccataggat
     3601 ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac aacctattaa
     3661 tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga cgactgaatc
     3721 cggtgagaat ggcaaaagct tatgcatttc tttccagact tgttcaacag gccagccatt
     3781 acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg
     3841 agcgaggcga aatacgcgat cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa
     3901 ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag gatattcttc
     3961 taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg catcatcagg
     4021 agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc agtttagtct
     4081 gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca gaaacaactc
     4141 tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc cgacattatc
     4201 gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc gcggcctgga
     4261 gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc
     4321 agacagtttt attgttcatg atgatatatt tttatcttgt gcaatgtaac atcagagatt
     4381 ttgagacaca acgtggcttt gttgaataaa tcgaactttt gctgagttga aggatcagtc
     4441 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     4501 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     4561 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     4621 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     4681 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     4741 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     4801 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     4861 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     4921 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     4981 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     5041 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     5101 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     5161 atgttctttc ctgcgttatc ccctgattct gtggataacc gtgcggccgc caattaatac
     5221 gactcactat agggcgaatt ggggcgcgcc attctagatt aaccctagaa agatagtctg
     5281 cgtaaaattg acgcatgcat tcttgaaata ttgctctctc tttctaaata gcgcgaatcc
     5341 gtcgctgtgc atttaggaca tctcagtcgc cgcttggagc tcccgtgagg cgtgcttgtc
     5401 aatgcggtaa gtgtcactga
//
