LOCUS       sequence-136552-        5151 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  template for N-terminal eGFP tag, constitutive Puromycin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5151)
  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 5151)
  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: pRB39
FEATURES             Location/Qualifiers
     source          1..5151
                     /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)"
     CDS             join(1790..1792,1793..1795,1796..2506)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1790..1792,1793..1795,1796..2506)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1835..1856)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2096..2115)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2443..2464
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(join(2983..2989,2990..3007,3008..3013))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    3028..3049
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3169..3186)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3340..3928)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3420..3439)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4099..4890,4891..4959))
                     /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     4722..4741
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4960..5064)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5132..5150
                     /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 tggtgagcaa
     1801 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
     1861 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
     1921 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
     1981 cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt
     2041 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
     2101 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     2161 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     2221 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt
     2281 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca
     2341 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac
     2401 ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     2461 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaaggaat tcggtacctc
     2521 tccagctgct ccaagcggcg gtactagtat ctttctagaa gatctcctac aatattctca
     2581 gctgccatgg aaaatcgatg ttcttctttt attctctcaa gattttcagg ctgtatatta
     2641 aaacttatat taagaactat gctaaccacc tcatcaggaa ccgttgtagg tggcgtgggt
     2701 tttcttggca atcgactctc atgaaaacta cgagctaaat attcaatatg ttcctcttga
     2761 ccaactttat tctgcatttt ttttgaacga ggtttagagc aagcttcagg aaactgagac
     2821 aggaatttta ttaaaaattt aaattttgaa gaaagttcag ggttaatagc atccattttt
     2881 tgctttgcaa gttcctcagc attcttaaca aaagacgtct cttttgacat gtttaaagtt
     2941 taaacctcct gtgtgaaatt attatccgct cataattcca cacattatac gagccggaag
     3001 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     3061 ctcactgcca attgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     3121 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     3181 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     3241 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     3301 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     3361 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     3421 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     3481 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     3541 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     3601 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     3661 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     3721 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     3781 gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     3841 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     3901 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     3961 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     4021 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     4081 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     4141 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     4201 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     4261 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     4321 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     4381 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     4441 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     4501 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     4561 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     4621 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     4681 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     4741 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     4801 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     4861 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     4921 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     4981 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     5041 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga
     5101 aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc c
//
