LOCUS       sequence-333293-        5336 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  RMCE vector for generating C-terminal T2A-GAL4-DBD fusion (split 
            GAL4).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5336)
  AUTHORS   Kondo S, Takahashi T, Yamagata N, Imanishi Y, Katow H, Hiramatsu S, 
            Lynn K, Abe A, Kumaraswamy A, Tanimoto H
  TITLE     Neurochemical Organization of the Drosophila Brain Visualized by 
            Endogenously Tagged Neurotransmitter Receptors.
  JOURNAL   Cell Rep. 2020 Jan 7;30(1):284-297.e5. doi: 
            10.1016/j.celrep.2019.12.018.
  PUBMED    31914394
REFERENCE   2  (bases 1 to 5336)
  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: 
            "10.1016/j.celrep.2019.12"; journalName: "Cell Rep"; date: 
            "2020-01-7- 7"; volume: "30"; issue: "1"; pages: "284-297.e5"
COMMENT     Sequence Label: pF3BGX-T2A-GAL4-DBD
FEATURES             Location/Qualifiers
     source          1..5336
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    join(1..31,32..586)
                     /label=P element 5' end
     protein_bind    614..647
                     /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)"
     CDS             join(898..900,901..903,904..1617)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(943..964)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1204..1223)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1551..1572
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1733..1814
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1770..1789)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1824..1843
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1861..1894
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     CDS             1906..1959
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2146..2586
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="DNA binding domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 DNA binding domain
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     protein_bind    complement(2598..2631)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2644..2677
                     /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)"
     misc_feature    join(2704..2905,2906..2936)
                     /label=P element 3' end
                     /note="P element 3' end"
     primer_bind     complement(3042..3059)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3213..3801)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3293..3312)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3972..4763,4764..4832))
                     /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     4595..4614
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4833..4937)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5100..5121)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 catgatgaaa taacataagg tggtcccgtc gatagccgaa gcttaccgaa gtatacactt
       61 aaattcagtg cacgtttgct tgttgagagg aaaggttgtg tgcggacgaa tttttttttg
      121 aaaacattaa cccttacgtg gaataaaaaa aaatgaaata ttgcaaattt tgctgcaaag
      181 ctgtgactgg agtaaaatta attcacgtgc cgaagtgtgc tattaagaga aaattgtggg
      241 agcagagcct tgggtgcagc cttggtgaaa actcccaaat ttgtgatacc cactttaatg
      301 attcgcagtg gaaggctgca cctgcaaaag gtcagacatt taaaaggagg cgactcaacg
      361 cagatgccgt acctagtaaa gtgatagagc ctgaaccaga aaagataaaa gaaggctata
      421 ccagtgggag tacacaaaca gagtaagttt gaatagtaaa aaaaatcatt tatgtaaaca
      481 ataacgtgac tgtgcgttag gtcctgttca ttgtttaatg aaaataagag cttgagggaa
      541 aaaattcgta ctttggagta cgaaatgcgt cgtttagagc agcagcggta ccgggccccc
      601 cctcgaggtc gacgaagttc ctattcttca aatagtatag gaacttcaag cttcttaagg
      661 aattcgagct cgcccgggga tctaattcaa ttagagacta attcaattag agctaattca
      721 attaggatca gcttatcgat ttcgaaccct cgaccgccgg agtataaata gaggcgcttc
      781 gtctacggag cgacaattca attcaaacaa gcaaagtgaa cacgtcgcta agcgaaagct
      841 aagcaaataa acaagcgcag ctgaacaagc taaacaatcg ggctagacct aggcaaaatg
      901 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
      961 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     1021 aagctgaccc tgaagctcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     1081 gtgaccaccc tgggctacgg cctgcagtgc ttcgcccgct accccgacca catgaagcag
     1141 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     1201 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     1261 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     1321 ctggagtaca actacaacag ccacaacgtc tatatcaccg ccgacaagca gaagaacggc
     1381 atcaaggcca acttcaagat ccgccacaac atcgaggacg gcggcgtgca gctcgccgac
     1441 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     1501 ctgagctacc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     1561 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaacca
     1621 tggctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct
     1681 cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt
     1741 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     1801 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cgaattcgta
     1861 cgcgcccggg gagcccaagg gcacgccctg gcacccggcg cgccagaggg ccgcggcagc
     1921 ctgctcacct gcggcgacgt ggaagagaac ccaggaccaa tgctggagat ccgcgccgcc
     1981 ttcctgcgcc agcgcaacac cgccctgcgc accgaggtgg ccgagctgga gcaggaggtg
     2041 cagcgcctgg agaacgaggt gagccagtac gagacccgct acggccccct gggcggcggc
     2101 aaggctagcg gaggaggtgg tggaggtgga ggtggaggta ctagtatgaa gctactgtct
     2161 tctatcgaac aagcatgcga tatttgccga cttaaaaagc tcaagtgctc caaagaaaaa
     2221 ccgaagtgcg ccaagtgtct gaagaacaac tgggagtgtc gctactctcc caaaaccaaa
     2281 aggtctccgc tgactagggc acatctgaca gaagtggaat caaggctaga aagactggaa
     2341 cagctatttc tactgatttt tcctcgagaa gaccttgaca tgattttgaa aatggattct
     2401 ttacaggata taaaagcatt gttaacagga ttatttgtac aagataatgt gaataaagat
     2461 gccgtcacag atagattggc ttcagtggag actgatatgc ctctaacatt gagacagcat
     2521 agaataagtg cgacatcatc atcggaagag agtagtaaca aaggtcaaag acagttgact
     2581 gtatcgtaag cggccgcata acttcgtata gcatacatta tacgaagtta taccggtggc
     2641 cgcgaagttc ctattcttca aatagtatag gaacttcgag ctcccggaga gatgcagaag
     2701 ctttgcgtac tcgcaaatta ttaaaaataa aactttaaaa ataatttcgt ctaattaata
     2761 ttatgagtta attcaaaccc cacggacatg ctaagggtta atcaacaatc atatcgctgt
     2821 ctcactcaga ctcaatacga cactcagaat actattcctt tcactcgcac ttattgcaag
     2881 catacgttaa gtggatgtct cttgccgacg ggaccacctt atgttatttc atcatggtct
     2941 ggccattctc atcgtgagct tccgggtgct cgccagctgc attaatgaat cggccaacgc
     3001 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     3061 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     3121 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     3181 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     3241 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     3301 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     3361 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     3421 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     3481 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     3541 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     3601 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     3661 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     3721 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     3781 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     3841 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     3901 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     3961 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     4021 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     4081 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     4141 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     4201 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     4261 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     4321 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     4381 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     4441 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     4501 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     4561 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     4621 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     4681 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     4741 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     4801 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     4861 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     4921 caaatagggg ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata
     4981 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     5041 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     5101 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     5161 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     5221 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     5281 ggggaaagcc ggccttctcc gccgccgatc tgccgctgga ctacgtgggt ctggcc
//
