LOCUS       sequence-397504-        3392 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tagging endogenously expressed proteins with HA epitope, C-terminal.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3392)
  AUTHORS   Gao Y, Hisey E, Bradshaw TWA, Erata E, Brown WE, Courtland JL, Uezu 
            A, Xiang Y, Diao Y, Soderling SH
  TITLE     Plug-and-Play Protein Modification Using Homology-Independent 
            Universal Genome Engineering.
  JOURNAL   Neuron. 2019 Aug 21;103(4):583-597.e8. doi: 
            10.1016/j.neuron.2019.05.047. Epub 2019 Jul 1.
  PUBMED    31272828
REFERENCE   2  (bases 1 to 3392)
  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"; doi: 
            "10.1016/j.neuron.2019.05.047"; journalName: "Neuron"; date: 
            "2019-08-21- 21"; volume: "103"; issue: "4"; pages: "583-597.e8"
COMMENT     Sequence Label: pHiUGE HA-epitope donor ORF0
FEATURES             Location/Qualifiers
     source          1..3392
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        145..393
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     145..165
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     316..335
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        414..489
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             532..558
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(574..600)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     repeat_region   655..795
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   655..784
                     /label=AAV2 ITR
     rep_origin      870..1325
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(957..976)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1167..1188
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1342..1361)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1461..1483
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1521..1539)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1607..1711
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1712..1780,1781..2572)
                     /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(1930..1949)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2743..3331
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3232..3251
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtgagggc ctatttccca tgattccttc atatttgcat
      181 atacgataca aggctgttag agagataatt ggaattaatt tgactgtaaa cacaaagata
      241 ttagtacaaa atacgtgacg tagaaagtaa taatttcttg ggtagtttgc agttttaaaa
      301 ttatgtttta aaatggacta tcatatgctt accgtaactt gaaagtattt cgatttcttg
      361 gctttatata tcttgtggaa aggacgaaac accgacgctt ccgagtacgg tacgttttag
      421 agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa agtggcaccg
      481 agtcggtgct ttttttctag acctgtaccg tactcggaag cgtcggatcc atacccttac
      541 gatgtaccgg attacgcata gctagctagc tagagcatag tcaggaacgt catagggata
      601 aagctcgagc ctgtaccgta ctcggaagcg tccacgtgcg gaccgagcgg ccgcaggaac
      661 ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc
      721 gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc
      781 gcagctgcct gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
      841 cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc
      901 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc
      961 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     1021 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     1081 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     1141 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     1201 ctctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg tctattggtt
     1261 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     1321 aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     1381 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     1441 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     1501 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     1561 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     1621 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     1681 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     1741 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     1801 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     1861 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     1921 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     1981 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     2041 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     2101 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     2161 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     2221 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     2281 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     2341 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     2401 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     2461 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     2521 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     2581 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     2641 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     2701 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     2761 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     2821 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     2881 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     2941 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     3001 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     3061 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     3121 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     3181 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     3241 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     3301 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     3361 cctggccttt tgctggcctt ttgctcacat gt
//
