LOCUS       sequence-494250-        8145 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Conditional tagging of endogenous proteins with smGFP-HA in open 
            reading frame 2, with dual membrane labelling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8145)
  TITLE     dFlpTag conditional protein labelling lines
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8145)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pFLPtag-SA-SD-2-smGFP-HA
FEATURES             Location/Qualifiers
     source          1..8145
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          1527..1592
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             1722..1742
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    2014..2148
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2051..2070)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2105..2124
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             3079..3105
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3109..3135
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3139..3165
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3694..3720
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3727..3753
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3766..3792
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3799..3825
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4036..4062
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4066..4092
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4096..4122
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    complement(4314..4347)
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     protein_bind    complement(4412..4459)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        complement(4489..4727)
                     /label=hsp70 promoter
                     /note="Drosophila melanogaster hsp70Bb promoter"
     primer_bind     complement(4521..4542)
                     /label=pCasper-hs
                     /note="Drosophila Hsp70 promoter, forward primer"
     protein_bind    4746..4840
                     /label=5X UAS
                     /bound_moiety="GAL4"
                     /note="five tandem copies of the ""ScaI site"" 17-mer 
                     CGGAGTACTGTCCTCCG, an upstream activating sequence (UAS) 
                     that efficiently binds yeast Gal4 (Webster et al., 1988; 
                     Pfeiffer et al., 2010)"
     protein_bind    4895..4964
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     primer_bind     complement(5004..5024)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5004..5022)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5043..5059)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5043..5059)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5056..5078)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5067..5083
                     /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)."
     promoter        complement(join(5091..5097,5098..5115,5116..5121))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5136..5157
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5274..5291)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5445..6033)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5525..5544)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6204..6995,6996..7064))
                     /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     6827..6846
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7065..7169)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(7195..7650)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7332..7353)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7544..7563
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7777..7799
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7791..7808
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7792..7808
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7818..7837
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7818..7836
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    7872..7941
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     protein_bind    8012..8045
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     protein_bind    8096..8143
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
ORIGIN
        1 ggccgctatg gtgtccaagg gcgaggaagt gatcaaggag ttcatgcgct tcaaagtgcg
       61 catggagggc agcatgaacg gccacgagtt cgagattgag ggcgagggcg agggacgccc
      121 atatgaggga acccagaccg ccaagctgaa agtgaccaag ggcggacccc tgcccttcgc
      181 ctgggatatt ctgagccccc agtttatgta cggcagcaag gcctacgtga agcaccccgc
      241 cgatatcccc gattacaaga agctgagctt ccccgagggc ttcaagtggg agcgcgtgat
      301 gaacttcgag gatggcggcc tcgtgaccgt gacccaggat agcagtctgc aggatggcac
      361 cctgatctac aaagtgaaga tgcgcggcac caacttcccc cccgatggac ccgtgatgca
      421 gaagaagacc atgggctggg aggccagcac cgagcgcctg tatccacgcg acggcgtgct
      481 gaagggcgag attcatcagg ccctgaagct gaaggacggc ggccactacc tggtggagtt
      541 caagaccatc tacatggcca agaagcccgt gcagctgccc ggctactact acgtggacac
      601 caagctggat atcaccagcc acaacgagga ttacaccatc gtggagcagt acgagcgcag
      661 cgagggccgc caccatctgt ttctgggaca cggaaccggc agcaccggaa gcggaagtag
      721 tggaaccgcc agcagcgagg ataacaacat ggccgtgatt aaggagttta tgcgctttaa
      781 agtgcgcatg gagggatcca tgaatggaca tgagtttgag atcgagggcg agggcgaggg
      841 acgcccctac gagggcacgc agacggccaa gctgaaagtg acgaagggcg gaccgctgcc
      901 gtttgcctgg gacatcctgt cgccgcagtt tatgtatggc tcgaaggcct atgtgaagca
      961 cccggccgac attccggact ataagaagct gtcgttccca gagggattca agtgggagcg
     1021 cgtgatgaat tttgaggacg gcggactcgt gaccgtgacg caggacagtt ccctgcagga
     1081 cggaacgctg atctataaag tgaagatgcg cggaacgaat ttcccgccgg acggcccagt
     1141 gatgcagaag aagacgatgg gatgggaggc ctcgaccgag cgcctgtacc cccgcgacgg
     1201 cgtgctgaag ggcgagatcc accaggccct gaagctgaag gatggcggac gctatctggt
     1261 ggagtttaag acgatctata tggccaagaa gccagtgcag ctgccgggat attactatgt
     1321 ggatacgaag ctggacatta cctcgcacaa tgaggactat acgattgtgg agcagtatga
     1381 gcgctcggag ggacgccatc acctgtttct gtacggcatg gatgagctgt ataagtagga
     1441 ggccggcctc gatctttgtg aaggaacctt acttctgtgg tgtgacataa ttggacaaac
     1501 tacctacaga gatttaaagc tctaaggtaa atataaaatt tttaagtgta taatgtgtta
     1561 aactactgat tctaattgtt tgtgtatttt agattccaac ctatggaact gatgaatggg
     1621 agcagtggtg gaatgccttt aatgaggaaa acctgttttg ctcagaagaa atgccatcta
     1681 gtgatgatga ggctactgct gactctcaac attctactcc tccaaaaaag aagagaaagg
     1741 tagaagaccc caaggacttt ccttcagaat tgctaagttt tttgagtcat gctgtgttta
     1801 gtaatagaac tcttgcttgc tttgctattt acaccacaaa ggaaaaagct gcactgctat
     1861 acaagaaaat tatggaaaaa tatttgatgt atagtgcctt gactagagat cataatcagc
     1921 cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac
     1981 ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt
     2041 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     2101 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggatctg cgtcgcgcca
     2161 cttcaacgct cgatgggagc gtcattggtg ggcggggtaa ccgtcgaaat cagtgtttac
     2221 gcttccaatc gcaacaaaaa attcactgca acactgaaaa gcatacgaaa acgatgaaga
     2281 ttgtacgaga aaccataaag tattttatcc acaaagacac gtatagcaga aaagccaagt
     2341 taactcggcg ataagttgtg tacacaagaa taaaatcggc cagattcagt gttgtcagaa
     2401 ataagaaaac cccactatgt ttttctttgc cttttctttc tcccagcgat cattcatttc
     2461 gtggtgaaag aacggggtca ttgcacggag tttcgactgc gggaaagcag agctgccgtt
     2521 cacttcgtct ataattagcg ctttctattt tccccgattc gggccgctgc tgcgcttttc
     2581 cgcctgctgt ttgtggcaag tgtagcagca ggctgtgcac gcagtgtggc atgcacttgg
     2641 ctttccaccg ttggtatcga ttctctggga cgatgagtca ttcctttcgg gaccacagca
     2701 taatcgttgc cagctcaccg aaatggtgac ttcatttctt aactgccgtc aagcatgcga
     2761 ttgtacatac atacatattt atatatgtac atatttatgt gactatggta ggtcgatata
     2821 atagcaatca acgcaagcaa atgtgtcagt cctgcttaca ggaacgattc tatttagtaa
     2881 ttttcgttgt ataaagtaat tatgtatgta tgtaagcccc ataaatctga aaaccggtag
     2941 tcgatccaac atggcgactt gtcccatccc cggcatgttt aaatatacta attattcttg
     3001 aactaatttt aatcaaccga tttatctctc ttccgcaggt cgggaggttc cggtggaagc
     3061 ggaggtagcg gcggatccta cccttatgat gtgcccgatt atgctggcta cccttatgat
     3121 gttcctgatt acgccggata tccgtatgac gtgccagact acgcgggagg tgtgagcaag
     3181 ggagaggagt tgtttaccgg cgtggtcccc atactggtgg agttggatgg cgacgttaat
     3241 ggacataaat tctcggtccg cggcgaggga gagggagacg ccaccaatgg caagctgacg
     3301 cttaagttca tttgcacgac tggaaaattg cctgtcccct ggccgaccct ggtcaccaca
     3361 ctgggcggag gagtgcagtg cttctcccgt tacccagacc acatgaagca gcacgatttc
     3421 ttcaagagcg caatgccgga gggatacgtg caagaacgga ctatttcctt caaggatgac
     3481 ggaacctata agacccgtgc ggaggtcaaa ttcgagggtg acaccctggt gaaccgaatt
     3541 gaactcaaag gaatcgattt caaggaggat ggaaatattc tgggtcacaa gctggaatac
     3601 aacttcaaca gccataatgt gtacattacg gctgataagc agaagaacgg cattaaggcc
     3661 aatttcaaga tccgccacaa cgttgagggt ggatatccct acgacgtgcc cgattatgcg
     3721 ggcggatatc cgtatgatgt tccagattat gctggtggtg gaggctatcc ctatgatgtc
     3781 cccgactacg ccggaggtta cccatacgat gtgcccgact acgctggagg cgatggcagc
     3841 gtgcagctgg cagatcatta tcaacagaat acccccatag gtgatggccc cgttctgctt
     3901 ccagataatc actacctttc cacccagagc gtgctttcga aagacccgaa cgaaaagcgt
     3961 gatcacatgg tcctgctgga gtttgtgacc gcggctggaa tcaccctggg tatggacgaa
     4021 ctctacaagg gtggctaccc ctacgatgtc ccggactacg ctggatatcc gtatgacgta
     4081 ccggactatg ccggttaccc gtacgatgtc ccggactacg ctcgatccgg aggtagcggt
     4141 ggaagcggag gttccggcag gtaagttatt gaacaatggc atcaaatgcc ttcatcatca
     4201 ctacccttta gcccttaaga ccccacaatg accttaccca ctcagagaaa aaagtaaata
     4261 tgaaagccca tttgaacttc taagcttgtc gacttaatta aatcgatcct agggaagttc
     4321 ctatactatt tgaagaatag gaacttcgga ataggaactt ctaagaggtt tcatattgct
     4381 aatagcagct acaatccagc taccattctg cgaagttcct atactttcta gagaatagga
     4441 acttcggaat aggaacttcg ctggtaccct cgagcccaat tccctattca gagttctctt
     4501 cttgtattca ataattactt cttggcagat ttcagtagtt gcagttgatt tacttggttg
     4561 ctggttactt ttaattgatt cactttaact tgcactttac tgcagattgt ttagcttgtt
     4621 cagctgcgct tgtttatttg cttagctttc gcttagcgac gtgttcactt tgcttgtttg
     4681 aattgaattg tcgctccgta gacgaagcgc ctctatttat actccggcgc tcgctagagt
     4741 ctccgctcgg aggacagtac tccgctcgga ggacagtact ccgctcggag gacagtactc
     4801 cgctcggagg acagtactcc gctcggagga cagtactccg acctgcaggg aattcctgca
     4861 gactagttct agagtcgacc atcatgatgg accagatggg tgaggtggag tacgcgcccg
     4921 gggagcccaa gggcacgccc tggcacccgc accgcggctt cgagaccgtg acctacatca
     4981 tgcatggtac ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
     5041 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     5101 gccgggagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     5161 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     5221 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     5281 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     5341 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     5401 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     5461 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     5521 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     5581 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     5641 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     5701 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     5761 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     5821 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     5881 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     5941 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     6001 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     6061 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     6121 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     6181 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     6241 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     6301 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     6361 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     6421 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     6481 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     6541 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     6601 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     6661 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     6721 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     6781 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     6841 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     6901 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     6961 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     7021 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     7081 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     7141 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg
     7201 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     7261 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     7321 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     7381 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     7441 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     7501 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     7561 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     7621 ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt
     7681 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
     7741 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
     7801 cggccagtga gcgcgcgtaa tacgactcac tatagggcga attggagctc gctagcgatg
     7861 taggtcacgg tctcgaagcc gcggtgcggg tgccagggcg tgcccttggg ctccccgggc
     7921 gcgtactcca cctcacccat ctggtccatc atgatggtcg acaagcttgg atccgatgtc
     7981 ccaatacgcg taggcctgaa gttcctattc cgaagttcct attcttcaaa tagtatagga
     8041 acttctcttt gcaccatttc aaagaataac agtgataatt tctgggttaa ggcaagaagt
     8101 tcctattccg aagttcctat tctctagaaa gtataggaac ttcgc
//
