LOCUS       sequence-136533-        4035 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  template for C-terminal double-FLAG tag, Puromycin-resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4035)
  AUTHORS   Kunzelmann S, Bottcher R, Schmidts I, Forstemann K
  TITLE     A Comprehensive Toolbox for Genome Editing in Cultured Drosophila 
            melanogaster Cells.
  JOURNAL   G3 (Bethesda). 2016 Jun 1;6(6):1777-85. doi: 10.1534/g3.116.028241.
  PUBMED    27172193
REFERENCE   2  (bases 1 to 4035)
  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: "10.1534/g3.116"; 
            journalName: "G3 (Bethesda)"; date: "2016-06-1- 1"; volume: "6"; 
            issue: "6"; pages: "1777-85"
COMMENT     Sequence Label: pSK25
FEATURES             Location/Qualifiers
     source          1..4035
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     305..324
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        305..323
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             358..381
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             391..414
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    complement(424..457)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     protein_bind    complement(1399..1432)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     promoter        complement(join(1867..1873,1874..1891,1892..1897))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    1912..1933
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2053..2070)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2224..2812)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2304..2323)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2983..3774,3775..3843))
                     /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     3606..3625
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3844..3948)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4016..4034
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcccctgcag ccgaattata ttatttttgc caaataattt ttaacaaaag ctctgaagtc
       61 ttcttcattt aaattcttag atgatacttc atctggaaaa ttgtcccaat tagtagcatc
      121 acgctgtgag taagttctaa accatttttt tattgttgta ttatctctaa tcttactact
      181 cgatgagttt tcggtattat ctctattttt aacttggagc aggttccatt cattgttttt
      241 ttcatcatag tgaataaaat caactgcttt aacacttgtg cctgaacacc atatccatcc
      301 ggcgtaatac gactcactat agggagagcg gccgccagat cttccggatg gctcgaggat
      361 tataaagatg atgatgataa atccggagcc gattataaag atgatgatga taaatgaact
      421 agtgaagttc ctatactttc tagagaatag gaacttcaaa taaacatatg ggcgcgccct
      481 gttggaatat actattcaac ctacaaaaat aacgttaaac aacactactt tatatttgat
      541 atgaatggcc acacctttta tgccataaaa catattgtaa gagaatacca ctctttttat
      601 tccttctttc cttcttgtac gttttttgct gtgagtaggt cgtggtgctg gtgttgcagt
      661 tgaaataact taaaatataa atcataaaac tcaaacataa acttgactat ttatttattt
      721 attaagaaag gaggtaccgg tggcgcgcca tgaccgagta caagcctaca gtgcgactgg
      781 ccacacgcga tgacgttccc agagccgttc ggacgttggc tgccgcgttt gcggactatc
      841 ccgcaactcg ccatactgtg gaccctgatc gccacatcga gagggtgact gagcttcagg
      901 agctgttcct gaccagggta gggctggaca taggcaaggt ctgggtggcc gacgatggag
      961 ccgctgttgc ggtgtggact accccagaga gcgtggaagc aggggctgtc tttgccgaaa
     1021 ttgggccaag gatggccgaa ctgtctgggt cccgtttggc tgcccaacag cagatggaag
     1081 gactgcttgc cccacatcga cccaaggagc ctgcatggtt cctcgctacc gtgggtgtga
     1141 gcccagatca ccagggcaaa ggtctgggat cagccgtcgt tctgcctggc gtcgaggctg
     1201 ctgagagagc aggagtaccg gcattcctcg aaaccagtgc tccacggaac ctcccgttct
     1261 acgagagact gggctttacg gtcacagccg atgtggaagt gcccgaaggc cctagaacct
     1321 ggtgcatgac acggaaaccc ggtgcatgaa aattaaatat aaataaagct attaaaataa
     1381 attactatct aacatatgga agttcctata ctttctagag aataggaact tcatctttct
     1441 agaagatctc ctacaatatt ctcagctgcc atggaaaatc gatgttcttc ttttattctc
     1501 tcaagatttt caggctgtat attaaaactt atattaagaa ctatgctaac cacctcatca
     1561 ggaaccgttg taggtggcgt gggttttctt ggcaatcgac tctcatgaaa actacgagct
     1621 aaatattcaa tatgttcctc ttgaccaact ttattctgca ttttttttga acgaggttta
     1681 gagcaagctt caggaaactg agacaggaat tttattaaaa atttaaattt tgaagaaagt
     1741 tcagggttaa tagcatccat tttttgcttt gcaagttcct cagcattctt aacaaaagac
     1801 gtctcttttg acatgtttaa agtttaaacc tcctgtgtga aattattatc cgctcataat
     1861 tccacacatt atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     1921 ctaactcaca ttaattgcgt tgcgctcact gccaattgct ttccagtcgg gaaacctgtc
     1981 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     2041 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     2101 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     2161 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     2221 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     2281 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     2341 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     2401 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     2461 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     2521 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     2581 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     2641 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt
     2701 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     2761 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     2821 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     2881 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     2941 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     3001 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     3061 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     3121 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     3181 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     3241 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     3301 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     3361 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     3421 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     3481 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     3541 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     3601 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     3661 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     3721 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     3781 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     3841 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     3901 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     3961 aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag
     4021 gcgtatcacg aggcc
//
