LOCUS       sequence-398255-        7099 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express ""G"" HaloTag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7099)
  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 7099)
  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: pJPT004-1x FLAG-HaloTag-G
FEATURES             Location/Qualifiers
     source          1..7099
                     /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    1082..1203
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1093..1112)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1147..1166
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1318..1364
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1394..1530
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1575..1646
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1872..2460)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1952..1971)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2960..4169
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4006..4025
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4173..4772
                     /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(4173..4192)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4669..4689
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4799..4847
                     /note="synthetic polyadenylation signal"
     CDS             join(5113..5181,5182..5973)
                     /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(5331..5350)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5986..6122
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6274..6345
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6350..6396)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6414..6462
                     /note="synthetic polyadenylation signal"
     enhancer        6543..6846
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6848..7050
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7001..7021
                     /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 gctttcgtag tgatggcaaa aagaaaaaga agaaatccaa gaccaaatgc
     1021 cagctgctgt gagctagctt gactgactga gatacagcgt accttcagct cacagacatg
     1081 ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
     1141 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     1201 gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt gtgggaggtt
     1261 ttttaaagca agtaaaacct ctacaaatgt ggtattggcc catctctatc ggtatcgtag
     1321 cataacccct tggggcctct aaacgggtct tgaggggttt tttgtgcccc tcgggccgga
     1381 ttgctatcta ccggcattgg cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata
     1441 ctcccacata tgccagattc agcaacggat acggcttccc caacttgccc acttccatac
     1501 gtgtcctcct taccagaaat ttatccttaa ggtcgtcagc tatcctgcag gcgatctctc
     1561 gatttcgatc aagacattcc tttaatggtc ttttctggac accactaggg gtcagaagta
     1621 gttcatcaaa ctttcttccc tccctaatct cattggttac cttgggctat cgaaacttaa
     1681 ttaaccagtc aagtcagcta cttggcgaga tcgacttgtc tgggtttcga ctacgctcag
     1741 aattgcgtca gtcaagttcg atctggtcct tgctattgca cccgttctcc gattacgagt
     1801 ttcatttaaa tcatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     1861 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     1921 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     1981 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     2041 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     2101 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     2161 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     2221 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     2281 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     2341 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     2401 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     2461 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     2521 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     2581 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     2641 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcattt
     2701 aaatttccga actctccaag gccctcgtcg gaaaatcttc aaacctttcg tccgatccat
     2761 cttgcaggct acctctcgaa cgaactatcg caagtctctt ggccggcctt gcgccttggc
     2821 tattgcttgg cagcgcctat cgccaggtat tactccaatc ccgaatatcc gagatcggga
     2881 tcacccgaga gaagttcaac ctacatcctc aatcccgatc tatccgagat ccgaggaata
     2941 tcgaaatcgg ggcgcgcctg gcctccgcgc cgggttttgg cgcctcccgc gggcgccccc
     3001 ctcgtcacgg cgagcgctgc cacgtcagac gaagggcgca ggagcgtcct gatccttccg
     3061 cccggacgct caggacagcg gcccgctgct cataagactc ggccttagaa ccccagtatc
     3121 agcagaagga cattttagga cgggacttgg gtgactctag ggcactggtt ttctttccag
     3181 agagcggaac aggcgaggaa aagtagtccc ttctcggcga ttctgcggag ggatctccgt
     3241 ggggcggtga acgccgatga ttatataagg acgcgccggg tgtggcacag ctagttccgt
     3301 cgcagccggg atttgggtcg cggttcttgt ttgtggatcg ctgtgatcgt cacttggtga
     3361 gtagcgggct gctgggctgg ccggggcttt cgtggccgcc gggccgctcg gtgggacgga
     3421 agcgtgtgga gagaccgcca agggctgtag tctgggtccg cgagcaaggt tgccctgaac
     3481 tgggggttgg ggggagcgca gcaaaatggc ggctgttccc gagtcttgaa tggaagacgc
     3541 ttgtgaggcg ggctgtgagg tcgttgaaac aaggtggggg gcatggtggg cggcaagaac
     3601 ccaaggtctt gagcccttcg ctaatgcggg aaagctctta ttcgggtgag atgggctggg
     3661 caccatctgg ggaccctgac gtgaagtttg tcactgactg gagaactcgg tttgtcgtct
     3721 gttgcggggg cggcagttat ggcggtgccg ttgggcagtg cacccgtacc tttgggagcg
     3781 cgcgccctcg tcgtgtcgtg acgtcacccg ttctgttggc ttataatgca gggtggggcc
     3841 acctgccggt aggtgtgcgg taggcttttc tccgtcgcag gacgcagggt tcgggcctag
     3901 ggtaggctct cctgaatcga caggcgccgg acctctggtg aggggaggga taagtgaggc
     3961 gtcagtttct ttggtcggtt ttatgtacct atcttcttaa gtagctgaag ctccggtttt
     4021 gaactatgcg ctcggggttg gcgagtgtgt tttgtgaagt tttttaggca ccttttgaaa
     4081 tgtaatcatt tgggtcaata tgtaattttc agtgttagac ttgtaaattg tccgctaaat
     4141 tctggccgtt tttggctttt ttgttagaca acatgaccga gtacaagccc acggtgcgcc
     4201 tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg ttcgccgact
     4261 accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc accgagctgc
     4321 aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg
     4381 gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg
     4441 agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg
     4501 agggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggcg
     4561 tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg
     4621 ccgccgagcg cgccggggtg cccgccttcc tggagacctc cgcgccccgc aacctcccct
     4681 tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca
     4741 cctggtgcat gacccgcaag cccggtgcct aggctagtat gtaagcctag tcttagataa
     4801 taaaatcgct atccatcgaa gatggatgtg tgttggtttt ttgtgtgtgt aacgctaggc
     4861 gcgcctggtg taccgagaac gatcctctca gtgcgagtct cgacgatcca tatcgttgct
     4921 tggcagtcag ccagtcggaa tccagcttgg gacccaggaa gtccaatcgt cagatattgt
     4981 actcaagcct ggtcacggca gcgtaccgat ctgtttaaac ctagatattg atagtctgat
     5041 cggtcaacgt ataatcgagt cctagctttt gcaaacatct atcaagagac aggatcagca
     5101 ggaggctttc gcatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     5161 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     5221 cagttgggtg cgcgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     5281 agttttcgcc ccgaagaacg ctttccaatg atgagcactt ttaaagttct gctatgtggc
     5341 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     5401 cagaatgact tggttgagta ttcaccagtc acagaaaagc atcttacgga tggcatgaca
     5461 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     5521 ctgacaacga ttggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     5581 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     5641 gacaccacga tgcctgtagc aatggcaaca accttgcgta aactattaac tggcgaacta
     5701 cttactctag cttcccggca acagttgata gactggatgg aggcggataa agttgcagga
     5761 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     5821 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     5881 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     5941 gagataggtg cctcactgat taagcattgg taaccgattc taggtgcatt ggcgcagaaa
     6001 aaaatgcctg atgcgacgct gcgcgtctta tactcccaca tatgccagat tcagcaacgg
     6061 atacggcttc cccaacttgc ccacttccat acgtgtcctc cttaccagaa atttatcctt
     6121 aagatcccga atcgtttaaa ctcgactctg gctctatcga atctccgtcg tttcgagctt
     6181 acgcgaacag ccgtggcgct catttgctcg tcgggcatcg aatctcgtca gctatcgtca
     6241 gcttaccttt ttggcagcga tcgcggctcc cgacatcttg gaccattagc tccacaggta
     6301 tcttcttccc tctagtggtc ataacagcag cttcagctac ctctcaattc aaaaaacccc
     6361 tcaagacccg tttagaggcc ccaaggggtt atgctatcaa tcgttgcgtt acacacacaa
     6421 aaaaccaaca cacatccatc ttcgatggat agcgatttta ttatctaact gctgatcgag
     6481 tgtagccaga tctagtaatc aattacgggg tcattagttc atagcccata tatggagttc
     6541 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     6601 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     6661 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
     6721 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
     6781 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
     6841 accatgctga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
     6901 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
     6961 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
     7021 gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat cagatctttg
     7081 tcgatcctac catccactc
//
