LOCUS       sequence-296119-        6016 bp    DNA     circular SYN 14-APR-2025
DEFINITION  a targeting plasmid for EGFP-PML.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6016)
  AUTHORS   Kurihara M, Kato K, Sanbo C, Shigenobu S, Ohkawa Y, Fuchigami T, 
            Miyanari Y
  TITLE     Genomic Profiling by ALaP-Seq Reveals Transcriptional Regulation by 
            PML Bodies through DNMT3A Exclusion.
  JOURNAL   Mol Cell. 2020 May 7;78(3):493-505.e8. doi: 
            10.1016/j.molcel.2020.04.004. Epub 2020 Apr 29.
  PUBMED    32353257
REFERENCE   2  (bases 1 to 6016)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.molcel.2020.04.004"; journalName: "Mol Cell"; date: 
            "2020-05-7- 7"; volume: "78"; issue: "3"; pages: "493-505.e8"
COMMENT     Sequence Label: pGb_PMLlxP_EGFP
FEATURES             Location/Qualifiers
     source          1..6016
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     49..71
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     63..80
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     64..80
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(1023..1056)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             1063..1659
                     /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(1063..1082)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1559..1579
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             1678..1734
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     protein_bind    complement(1741..1774)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             join(1795..1797,1798..1800,1801..2511)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1795..1797,1798..1800,1801..2511)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1840..1861)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2101..2120)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2448..2469
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2524..2550
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4009..4800
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      4893..5481
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5382..5401
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5635..5652
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(5811..5838)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(5930..6016)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 atgcctggca gttccctact ctcgcgttaa cgctagcatg gatgttttcc cagtcacgac
       61 gttgtaaaac gacggccagt cttaagcgtc tccccctgaa cctgaccccg gaccaagtgg
      121 tgggaattca tggcgagggc gatgccacct acggcagacc cattcaacgc tgttctgtca
      181 ccacggtcct tccactgact cagaactaac tctgtgtccc acagttactt ctcataggcc
      241 acctgcctca aggtcattca agagacgcag ctgacacgct gtaagaaacc cttcaccaac
      301 tctcagatca tatgtagaat cggtccattt ctcaccgtat gattcatctc atcttctctg
      361 gtatcacttc gcaggagccc catcttaaat ctgaaaatgg ccctgactac ttcccaaact
      421 ggctctcctg tttcagacct ccactccttt ccaagtgttt gtctctcttt aaaatgtctg
      481 agcagctctg aattcctggt cattttctat ctcatgctcc attctcagct gagggatgcc
      541 tttccaggca aacccatccc agtactgagt cgaagtctcc actagggtgc acttgggagg
      601 ctctctcaaa gaggagttta tttctctacc tggcttttaa ctagaaagac atatagctgt
      661 ggaatgagtc caatagctca cagacaggga aaagcctgtc agctctgttt ctctcctggt
      721 gtgcatgctg cgtacgtggc cggatctttc cttgttctgc taatgagcca tgcccgcatg
      781 ccgggcttgt ttacttgctt gtttggttaa cctttctgac tctaccagag ttacttccat
      841 agctccctag tctttctctt aactttggaa gggagattat agaaccagcc tctccgcccc
      901 tagacccgcc cagccctgtc tcacttcctc tcgggaggcg ggctctcctc gggcaatctc
      961 tcgctcagtt ttctggctca gatctgagac ccatttcccg ccttcacaga cctaatcaag
     1021 aaataacttc gtatagcata cattatacga agttatctcg agatgaccga gtacaagccc
     1081 acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg
     1141 ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc
     1201 accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc
     1261 gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg
     1321 gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag
     1381 caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc
     1441 accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc
     1501 ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc cgcgccccgc
     1561 aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa
     1621 ggaccgcgca cctggtgcat gacccgcaag cccggtgcca gcggccgcgg aagcggagct
     1681 actaacttca gcctgctgaa gcaggctgga gacgtggagg agaaccctgg accttctaga
     1741 ataacttcgt atagcataca ttatacgaag ttattggcta gcgccaccga gctcatggtg
     1801 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     1861 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gagacgctac gtacggcaag
     1921 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     1981 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     2041 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     2101 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     2161 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     2221 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
     2281 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     2341 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     2401 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     2461 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa ggatatcgaa
     2521 ttctacccat acgatgttcc agattacgct ggtggaggcg gaggttctgg gggaggaggt
     2581 agtggcggtg gtggttcagg aggcggcgga agcttggatc caggtggagg tggaagcggt
     2641 atggaaactg aaccagtttc cgtgcagaag gtacctgcac cccctggatc tccctgtcga
     2701 caacaggact ctgccctgac ccccacgccc accatgcctc ccccagagga accctccgaa
     2761 gactatgaac acagccaaag ccctgcagag gtactactgg gttagaaggt gtagggctat
     2821 taggtttttt tattcgttta ctttggtcga ggagttgagg atgaagaagg tcgttataga
     2881 gaagtaagtt aatgggttag tgtcctaaag agacaccaag gtcctgaccc tcctggctgg
     2941 gcagaggtag aactgatagc ctagagtaaa ataggaacag gggcccggcg gtgtggggca
     3001 cacctctaat cccaacactt gggaggcaga ggcaggtgga tttctgagtt caaggccagc
     3061 ctggtctcta cagagtgagt tccaggacag ccagggctac acagagaaac cctgtctaaa
     3121 aaaaaaaaaa aaaaaaccgg gatgagttgg attgtaactg gggatagttg gattgtaact
     3181 cctactccct gaagccttgc ctgaagcgcc tacctcagat gaaagactat agtcctgtgg
     3241 aaggcctggc caggagtctt atttgctctg gcctggagct aaaggcatgt cccagacttg
     3301 gcaaaactat agtcgccctc cctgaccccc acaccaacac ttcccgtttt tcagctgtgg
     3361 ttgcacatcc caggagcaat agtgcttggg atggtagggc tggagggagg atgacaagtt
     3421 ccaggccagc ctaggccaca aagagagttc aagaccgctg agggcatcac agcaagacct
     3481 gtttcaaaac aaaacaaaca aacagacaga acctggctca gaccaaagtg cctgctaaga
     3541 ccaacaggat tcacacagct ccgtaggtgg catcgccctc gccgcaagca agcgctcgaa
     3601 acggtgcagc ggctgttgcc ggtgctgtgc caggaccatg gccgagacgt ctagaccagc
     3661 caggacagaa atgcctcgac ttcgctgcta cccaaggttg ccgggtgacg cacaccgtgg
     3721 aaacggatga aggcacgaac ccagtggaca taagcctgtt cggttcgtaa gctgtaatgc
     3781 aagtagcgta tgcgctcacg caactggtcc agaaccttga ccgaacgcag cggtggtaac
     3841 ggcgcagtgg cggttttcat ggcttgttat gactgttttt ttggggtaca gtctatgcct
     3901 cgggcatcca agcagcaagc gcgttacgcc gtgggtcgat gtttgatgtt atggagcagc
     3961 aacgatgtta cgcagcaggg cagtcgccct aaaacaaagt taaacattat gagggaagcg
     4021 gtgatcgccg aagtatcgac tcaactatca gaggtagttg gcgtcatcga gcgccatctc
     4081 gaaccgacgt tgctggccgt acatttgtac ggctccgcag tggatggcgg cctgaagcca
     4141 cacagtgata ttgatttgct ggttacggtg accgtaaggc ttgatgaaac aacgcggcga
     4201 gctttgatca acgacctttt ggaaacttcg gcttcccctg gagagagcga gattctccgc
     4261 gctgtagaag tcaccattgt tgtgcacgac gacatcattc cgtggcgtta tccagctaag
     4321 cgcgaactgc aatttggaga atggcagcgc aatgacattc ttgcaggtat cttcgagcca
     4381 gccacgatcg acattgatct ggctatcttg ctgacaaaag caagagaaca tagcgttgcc
     4441 ttggtaggtc cagcggcgga ggaactcttt gatccggttc ctgaacagga tctatttgag
     4501 gcgctaaatg aaaccttaac gctatggaac tcgccgcccg actgggctgg cgatgagcga
     4561 aatgtagtgc ttacgttgtc ccgcatttgg tacagcgcag taaccggcaa aatcgcgccg
     4621 aaggatgtcg ctgccgactg ggcaatggag cgcctgccgg cccagtatca gcccgtcata
     4681 cttgaagcta gacaggctta tcttggacaa gaagaagatc gcttggcctc gcgcgcagat
     4741 cagttggaag aatttgtcca ctacgtgaaa ggcgagatca ccaaggtagt cggcaaataa
     4801 ccctcgagcc acccatgacc aaaatccctt aacgtgagtt acgcgtcgtt ccactgagcg
     4861 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     4921 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     4981 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     5041 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     5101 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     5161 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     5221 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     5281 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     5341 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     5401 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     5461 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     5521 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     5581 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     5641 tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg
     5701 ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc
     5761 aacgcaatta atacgcgtac cgctagccag gaagagtttg tagaaacgca aaaaggccat
     5821 ccgtcaggat ggccttctgc ttagtttgat gcctggcagt ttatggcggg cgtcctgccc
     5881 gccaccctcc gggccgttgc ttcacaacgt tcaaatccgc tcccggcgga tttgtcctac
     5941 tcaggagagc gttcaccgac aaacaacaga taaaacgaaa ggcccagtct tccgactgag
     6001 cctttcgttt tatttg
//
