LOCUS       sequence-136549-        4503 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  template for N-terminal 3x FLAG tag, constitutive Puromycin 
            resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4503)
  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 4503)
  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: pRB33
FEATURES             Location/Qualifiers
     source          1..4503
                     /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"
     protein_bind    complement(372..404)
                     /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(1738..1771)
                     /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(2335..2341,2342..2359,2360..2365))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    2380..2401
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2521..2538)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2692..3280)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2772..2791)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3451..4242,4243..4311))
                     /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     4074..4093
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4312..4416)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4484..4502
                     /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 gctcgagttt
      361 ttcagcaaga taagttccta tactttctag agaataggaa cttccatatg ggcgcgccct
      421 gttggaatat actattcaac ctacaaaaat aacgttaaac aacactactt tatatttgat
      481 atgaatggcc acacctttta tgccataaaa catattgtaa gagaatacca ctctttttat
      541 tccttctttc cttcttgtac gttttttgct gtgagtaggt cgtggtgctg gtgttgcagt
      601 tgaaataact taaaatataa atcataaaac tcaaacataa acttgactat ttatttattt
      661 attaagaaag gaggtaccgg tggcgcgcca tgaccgagta caagcctaca gtgcgactgg
      721 ccacacgcga tgacgttccc agagccgttc ggacgttggc tgccgcgttt gcggactatc
      781 ccgcaactcg ccatactgtg gaccctgatc gccacatcga gagggtgact gagcttcagg
      841 agctgttcct gaccagggta gggctggaca taggcaaggt ctgggtggcc gacgatggag
      901 ccgctgttgc ggtgtggact accccagaga gcgtggaagc aggggctgtc tttgccgaaa
      961 ttgggccaag gatggccgaa ctgtctgggt cccgtttggc tgcccaacag cagatggaag
     1021 gactgcttgc cccacatcga cccaaggagc ctgcatggtt cctcgctacc gtgggtgtga
     1081 gcccagatca ccagggcaaa ggtctgggat cagccgtcgt tctgcctggc gtcgaggctg
     1141 ctgagagagc aggagtaccg gcattcctcg aaaccagtgc tccacggaac ctcccgttct
     1201 acgagagact gggctttacg gtcacagccg atgtggaagt gcccgaaggc cctagaacct
     1261 ggtgcatgac acggaaaccc ggtgcatgaa aattaaatat aaataaagct attaaaataa
     1321 attactatct aacatatgcc taggcttgtt gatttgtttt ttgtttattt actgcggtaa
     1381 ttcctagtct tagaagctta ttgaacttgt tgatgccttc tggttgggaa atcctcctct
     1441 ttatacccga tccagtccaa tcgaaactgt gggccgtggt gcccgataag ataagaaagc
     1501 cgtgtgcaag tcgatcggcg tgcgcaaaag cataatgctt tttctgcccg gcgatgataa
     1561 gcgatttgca cacgccctga taacaggaat tgattttttt ggtgcgcaaa gcagtgaaag
     1621 ctctataaga atgctggacc gccagtcgac aatcatcaaa cagtgctagc taacggctaa
     1681 gtgaatcaag tttattttaa tataatcaac aacaaaatat tttaacacaa acctagggaa
     1741 gttcctatac tttctagaga ataggaactt ctaatccaaa atgggatcca ccggtgaatt
     1801 cgactacaag gaccacgacg gcgactacaa ggaccacgac atcgactaca aggacgacga
     1861 caagggtacc tctccagctg ctccaagcgg cggtactagt atctttctag aagatctcct
     1921 acaatattct cagctgccat ggaaaatcga tgttcttctt ttattctctc aagattttca
     1981 ggctgtatat taaaacttat attaagaact atgctaacca cctcatcagg aaccgttgta
     2041 ggtggcgtgg gttttcttgg caatcgactc tcatgaaaac tacgagctaa atattcaata
     2101 tgttcctctt gaccaacttt attctgcatt ttttttgaac gaggtttaga gcaagcttca
     2161 ggaaactgag acaggaattt tattaaaaat ttaaattttg aagaaagttc agggttaata
     2221 gcatccattt tttgctttgc aagttcctca gcattcttaa caaaagacgt ctcttttgac
     2281 atgtttaaag tttaaacctc ctgtgtgaaa ttattatccg ctcataattc cacacattat
     2341 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     2401 aattgcgttg cgctcactgc caattgcttt ccagtcggga aacctgtcgt gccagctgca
     2461 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     2521 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     2581 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     2641 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     2701 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     2761 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     2821 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     2881 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     2941 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     3001 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     3061 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     3121 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     3181 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     3241 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     3301 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     3361 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     3421 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     3481 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     3541 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     3601 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     3661 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     3721 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     3781 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     3841 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     3901 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     3961 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     4021 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     4081 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     4141 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     4201 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     4261 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     4321 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     4381 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     4441 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     4501 gcc
//
