LOCUS       sequence-362375-        4573 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor plasmid to endogenously tag VIR gene in Drosophila 
            melanogaster ovarian somatic sheet (OSS) cells. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4573)
  AUTHORS   Meng Q, Stoyko D, Andrews CM, Konstantinidou P, Genzor P, O T, 
            Elchert AR, Benner L, Sobti S, Katz EY, Haase AD
  TITLE     Functional editing of endogenous genes through rapid selection of 
            cell pools (Rapid generation of endogenously tagged genes in 
            Drosophila ovarian somatic sheath cells).
  JOURNAL   Nucleic Acids Res. 2022 May 27. pii: 6594084. doi: 
            10.1093/nar/gkac448.
  PUBMED    35639929
REFERENCE   2  (bases 1 to 4573)
  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: "Nucleic 
            Acids Res. 2022 May 27. pii: 6594084. doi: 10.1093/nar/gkac448."
COMMENT     Sequence Label: Donor_C_FH_VIR
FEATURES             Location/Qualifiers
     source          1..4573
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     118..137
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     371..388
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    505..526
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(541..546,547..564,565..571)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    579..595
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     584..606
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     603..619
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     603..619
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1161..1226
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1245..1271
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1281..1337
                     /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"
     CDS             1338..1937
                     /codon_start=1
                     /gene="pac from Streptomyces"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVECPKDRATWCMTRKPGA"
     primer_bind     complement(1338..1357)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1834..1854
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(2576..2593)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2576..2592)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2585..2607)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2801..2820)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2920..2942
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2980..2998)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3066..3170
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3171..3239,3240..4031)
                     /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(3389..3408)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(4202..4573,1..217)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
       61 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
      121 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
      181 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
      241 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
      301 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
      361 gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc
      421 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
      481 agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc
      541 tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca
      601 cacaggaaac agctatgacc atgattacgc caagcttgca tgcctgcagg agaagatgtt
      661 tgtgtcgtcg gtaaggggtc gtggttttgc tcggccacca ccttccaggg gcgatttgtt
      721 ccgctcccgg ccgcccaaca cctcgcgacc gccatcgctg cacgtggacg acttcctggc
      781 cttggaaact tgtggcgccc agcccacagg acccaccggc tacaacaaga taccctcaat
      841 gctgagaggc tcccgggtgg gtcgcaaccg aggatctcga atctcggcag cagctgcctt
      901 taggtaaagt gtcacgatta ccacttagca ggataacgct aatgggctct ttttactctt
      961 ttaacaggca aaagaagatg atgcgcattg gttcgccctc cagctgggca gagtcccccg
     1021 gatcgtatcg gtctgcctcg gactcccact tcagtagcag cgactcgcac tattcgtcgc
     1081 cccactacag tggacggccg cgtggacgag gcctgcgttc caggccatcc tacctgagaa
     1141 ccagcggagg aggttccggt gactacaagg accacgacgg cgattataag gatcacgaca
     1201 tcgactacaa agacgacgat gacaaggctg gcgccagcag cgcctaccca tacgacgtac
     1261 cagattacgc tgctggatct gccacgaact tctctctgtt aaagcaagca ggagacgtgg
     1321 aagaaaaccc cggtcctatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg
     1381 acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc
     1441 acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca
     1501 cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg
     1561 tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca
     1621 tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc
     1681 cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc
     1741 agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg
     1801 gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg
     1861 gcttcaccgt caccgccgac gtcgagtgcc cgaaggaccg cgcgacctgg tgcatgaccc
     1921 gcaagcccgg tgcctgatag aggacattgc agcccaaccc gcttctcact gacacttccc
     1981 gtaaactcgg caaacgaacc agaactagat cctaagatgc ccgctgggca gctgaattcc
     2041 aatacttaat agtgtgtgtt gctttcgtat tagttcgctt tacttcccac tttgtatgta
     2101 agccaaaagt tagttttccg taattacact gtgcaaacgg acgtcgtgcg actccatggg
     2161 caactcctcg ttgctgacta tctgctcgtt gagcgacagt gccatcagcg agatcttctt
     2221 gtgcggatac ttctccgcgt gctgcttgag gaacttgtcg tagtagccca tgccatggcc
     2281 catccgagct ccgcagcggg tgaaggccac accgggcaca atgaagagat cgatgccgtg
     2341 ccctgtggtg atcgagtgtg tgtgtgtggg tttagaagga gctctgatgt atgcctagcc
     2401 actcaccgtt ggtcatggca tcctcgcgtg cctccttgaa gtccggctgc tttatgttcc
     2461 acttggtcag aggcaggctc tcgtactcct ccatgccgcg cagccgcacc atcttcatcc
     2521 tgctgccctc gtaggtgggc acaaagaagg atccccgggt accgagctcg aattcactgg
     2581 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
     2641 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
     2701 cccaacagtt gcgcagcctg aatggcgaat ggcgcctgat gcggtatttt ctccttacgc
     2761 atctgtgcgg tatttcacac cgcatatggt gcactctcag tacaatctgc tctgatgccg
     2821 catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
     2881 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     2941 ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt
     3001 tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa
     3061 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     3121 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     3181 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     3241 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     3301 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     3361 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     3421 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     3481 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     3541 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     3601 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     3661 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     3721 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     3781 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     3841 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     3901 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     3961 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     4021 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     4081 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     4141 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     4201 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     4261 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     4321 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     4381 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     4441 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     4501 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     4561 cctacaccga act
//
