LOCUS       sequence-398263-        7078 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express ""PPF"" HaloTag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7078)
  AUTHORS   Peruzzi JA, Gunnels TF, Edelstein HI, Lu P, Baker D, Leonard JN, 
            Kamat NP
  TITLE     Enhancing extracellular vesicle cargo loading and functional 
            delivery by engineering protein-lipid interactions.
  JOURNAL   Nat Commun. 2024 Jul 4;15(1):5618. doi: 10.1038/s41467-024-49678-z.
  PUBMED    38965227
REFERENCE   2  (bases 1 to 7078)
  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: "Nat 
            Commun."; date: "2024-07-4"; volume: "15(1):5618. doi"; pages: " 
            10.1038/s41467-024-49678-z"
COMMENT     Sequence Label: pJPT006-1x FLAG-HaloTag-PPF
FEATURES             Location/Qualifiers
     source          1..7078
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      22..31
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             31..54
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             70..954
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     polyA_signal    1061..1182
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1072..1091)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1126..1145
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1297..1343
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1373..1509
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1554..1625
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1851..2439)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1931..1950)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2939..4148
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3985..4004
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4152..4751
                     /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(4152..4171)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4648..4668
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4778..4826
                     /note="synthetic polyadenylation signal"
     CDS             join(5092..5160,5161..5952)
                     /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     complement(5310..5329)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5965..6101
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6253..6324
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6329..6375)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6393..6441
                     /note="synthetic polyadenylation signal"
     enhancer        6522..6825
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6827..7029
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6980..7000
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 gacacacccg ccagcggccg cgccaccatg gactacaagg acgatgacga caaggaattc
       61 ggcggctccg aaatcggtac tggctttcca ttcgaccccc attatgtgga agtcctgggc
      121 gagcgcatgc actacgtcga tgttggtccg cgcgatggca cccctgtgct gttcctgcac
      181 ggtaacccga cctcctccta cgtgtggcgc aacatcatcc cgcatgttgc accgacccat
      241 cgctgcattg ctccagacct gatcggtatg ggcaaatccg acaaaccaga cctgggttat
      301 ttcttcgacg accacgtccg cttcatggat gccttcatcg aagccctggg tctggaagag
      361 gtcgtcctgg tcattcacga ctggggctcc gctctgggtt tccactgggc caagcgcaat
      421 ccagagcgcg tcaaaggtat tgcatttatg gagttcatcc gccctatccc gacctgggac
      481 gaatggccag aatttgcccg cgagaccttc caggccttcc gcaccaccga cgtcggccgc
      541 aagctgatca tcgatcagaa cgtttttatc gagggtacgc tgccgatggg tgtcgtccgc
      601 ccgctgactg aagtcgagat ggaccattac cgcgagccgt tcctgaatcc tgttgaccgc
      661 gagccactgt ggcgcttccc aaacgagctg ccaatcgccg gtgagccagc gaacatcgtc
      721 gcgctggtcg aagaatacat ggactggctg caccagtccc ctgtcccgaa gctgctgttc
      781 tggggcaccc caggcgttct gatcccaccg gccgaagccg ctcgcctggc caaaagcctg
      841 cctaactgca aggctgtgga catcggcccg ggtctgaatc tgctgcaaga agacaacccg
      901 gacctgatcg gcagcgagat cgcgcgctgg ctgtctactc tggagatttc cggctccgga
      961 tccggtggta gcatgaaaaa gcatcgttgt aaatgttgca gcattatgtg agctagcttg
     1021 actgactgag atacagcgta ccttcagctc acagacatga taagatacat tgatgagttt
     1081 ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct
     1141 attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt
     1201 cattttatgt ttcaggttca gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc
     1261 tacaaatgtg gtattggccc atctctatcg gtatcgtagc ataacccctt ggggcctcta
     1321 aacgggtctt gaggggtttt ttgtgcccct cgggccggat tgctatctac cggcattggc
     1381 gcagaaaaaa atgcctgatg cgacgctgcg cgtcttatac tcccacatat gccagattca
     1441 gcaacggata cggcttcccc aacttgccca cttccatacg tgtcctcctt accagaaatt
     1501 tatccttaag gtcgtcagct atcctgcagg cgatctctcg atttcgatca agacattcct
     1561 ttaatggtct tttctggaca ccactagggg tcagaagtag ttcatcaaac tttcttccct
     1621 ccctaatctc attggttacc ttgggctatc gaaacttaat taaccagtca agtcagctac
     1681 ttggcgagat cgacttgtct gggtttcgac tacgctcaga attgcgtcag tcaagttcga
     1741 tctggtcctt gctattgcac ccgttctccg attacgagtt tcatttaaat catgtgagca
     1801 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     1861 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     1921 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     1981 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     2041 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2101 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     2161 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     2221 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     2281 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     2341 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     2401 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     2461 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     2521 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     2581 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     2641 tcagcgatct gtctatttcg ttcatccata gttgcattta aatttccgaa ctctccaagg
     2701 ccctcgtcgg aaaatcttca aacctttcgt ccgatccatc ttgcaggcta cctctcgaac
     2761 gaactatcgc aagtctcttg gccggccttg cgccttggct attgcttggc agcgcctatc
     2821 gccaggtatt actccaatcc cgaatatccg agatcgggat cacccgagag aagttcaacc
     2881 tacatcctca atcccgatct atccgagatc cgaggaatat cgaaatcggg gcgcgcctgg
     2941 cctccgcgcc gggttttggc gcctcccgcg ggcgcccccc tcgtcacggc gagcgctgcc
     3001 acgtcagacg aagggcgcag gagcgtcctg atccttccgc ccggacgctc aggacagcgg
     3061 cccgctgctc ataagactcg gccttagaac cccagtatca gcagaaggac attttaggac
     3121 gggacttggg tgactctagg gcactggttt tctttccaga gagcggaaca ggcgaggaaa
     3181 agtagtccct tctcggcgat tctgcggagg gatctccgtg gggcggtgaa cgccgatgat
     3241 tatataagga cgcgccgggt gtggcacagc tagttccgtc gcagccggga tttgggtcgc
     3301 ggttcttgtt tgtggatcgc tgtgatcgtc acttggtgag tagcgggctg ctgggctggc
     3361 cggggctttc gtggccgccg ggccgctcgg tgggacggaa gcgtgtggag agaccgccaa
     3421 gggctgtagt ctgggtccgc gagcaaggtt gccctgaact gggggttggg gggagcgcag
     3481 caaaatggcg gctgttcccg agtcttgaat ggaagacgct tgtgaggcgg gctgtgaggt
     3541 cgttgaaaca aggtgggggg catggtgggc ggcaagaacc caaggtcttg agcccttcgc
     3601 taatgcggga aagctcttat tcgggtgaga tgggctgggc accatctggg gaccctgacg
     3661 tgaagtttgt cactgactgg agaactcggt ttgtcgtctg ttgcgggggc ggcagttatg
     3721 gcggtgccgt tgggcagtgc acccgtacct ttgggagcgc gcgccctcgt cgtgtcgtga
     3781 cgtcacccgt tctgttggct tataatgcag ggtggggcca cctgccggta ggtgtgcggt
     3841 aggcttttct ccgtcgcagg acgcagggtt cgggcctagg gtaggctctc ctgaatcgac
     3901 aggcgccgga cctctggtga ggggagggat aagtgaggcg tcagtttctt tggtcggttt
     3961 tatgtaccta tcttcttaag tagctgaagc tccggttttg aactatgcgc tcggggttgg
     4021 cgagtgtgtt ttgtgaagtt ttttaggcac cttttgaaat gtaatcattt gggtcaatat
     4081 gtaattttca gtgttagact tgtaaattgt ccgctaaatt ctggccgttt ttggcttttt
     4141 tgttagacaa catgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc
     4201 ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg
     4261 tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg
     4321 tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga
     4381 ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg
     4441 agttgagcgg ttcccggctg gccgcgcagc aacagatgga gggcctcctg gcgccgcacc
     4501 ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca
     4561 agggtctggg cagcgccgtc gtgctccccg gagtggaggc cgccgagcgc gccggggtgc
     4621 ccgccttcct ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca
     4681 ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc
     4741 ccggtgccta ggctagtatg taagcctagt cttagataat aaaatcgcta tccatcgaag
     4801 atggatgtgt gttggttttt tgtgtgtgta acgctaggcg cgcctggtgt accgagaacg
     4861 atcctctcag tgcgagtctc gacgatccat atcgttgctt ggcagtcagc cagtcggaat
     4921 ccagcttggg acccaggaag tccaatcgtc agatattgta ctcaagcctg gtcacggcag
     4981 cgtaccgatc tgtttaaacc tagatattga tagtctgatc ggtcaacgta taatcgagtc
     5041 ctagcttttg caaacatcta tcaagagaca ggatcagcag gaggctttcg catgagtatt
     5101 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     5161 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc gcgagtgggt
     5221 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgc
     5281 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     5341 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtat
     5401 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     5461 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat tggaggaccg
     5521 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     5581 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     5641 atggcaacaa ccttgcgtaa actattaact ggcgaactac ttactctagc ttcccggcaa
     5701 cagttgatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     5761 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     5821 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     5881 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     5941 aagcattggt aaccgattct aggtgcattg gcgcagaaaa aaatgcctga tgcgacgctg
     6001 cgcgtcttat actcccacat atgccagatt cagcaacgga tacggcttcc ccaacttgcc
     6061 cacttccata cgtgtcctcc ttaccagaaa tttatcctta agatcccgaa tcgtttaaac
     6121 tcgactctgg ctctatcgaa tctccgtcgt ttcgagctta cgcgaacagc cgtggcgctc
     6181 atttgctcgt cgggcatcga atctcgtcag ctatcgtcag cttacctttt tggcagcgat
     6241 cgcggctccc gacatcttgg accattagct ccacaggtat cttcttccct ctagtggtca
     6301 taacagcagc ttcagctacc tctcaattca aaaaacccct caagacccgt ttagaggccc
     6361 caaggggtta tgctatcaat cgttgcgtta cacacacaaa aaaccaacac acatccatct
     6421 tcgatggata gcgattttat tatctaactg ctgatcgagt gtagccagat ctagtaatca
     6481 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
     6541 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
     6601 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
     6661 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     6721 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     6781 cctacttggc agtacatcta cgtattagtc atcgctatta ccatgctgat gcggttttgg
     6841 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
     6901 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
     6961 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
     7021 agcagagctg gtttagtgaa ccgtcagatc agatctttgt cgatcctacc atccactc
//
