LOCUS       sequence-275237-        6131 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  RMCE vector for generating C-terminal T2A-lexA-p65 fusion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6131)
  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 6131)
  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-lexAp65
FEATURES             Location/Qualifiers
     source          1..6131
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    28..61
                     /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(312..314,315..317,318..1031)
                     /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(357..378)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(618..637)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     965..986
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1147..1228
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1184..1203)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1238..1257
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1275..1308
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     CDS             1320..1373
                     /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             join(1374..1625,1626..1979)
                     /codon_start=1
                     /gene="lexA"
                     /product="E. coli DNA-binding protein that represses genes 
                     involved in the SOS response to DNA damage"
                     /label=LexA
                     /note="The N-terminal DNA-binding domain recognizes the 
                     16-bp palindromic sequence 5'-CTGTATATATATACAG-3'."
                     /translation="MKALTARQQEVFDLIRDHISQTGMPPTRAEIAQRLGFRSPNAAEE
                     HLKALARKGVIEIVSGASRGIRLLQEEEEGLPLVGRVAAGEPLLAQQHIEGHYQVDPSL
                     FKPNADFLLRVSGMSMKDIGIMDGDLLAVHKTQDVRNGQVVVARIDDEVTVKRLKKQGN
                     KVELLPENSEFKPIVVDLRQQSFTIEGLAVGVIRNGDWL"
     CDS             2424..2795
                     /codon_start=1
                     /product="transcriptional activation domain of human RelA, 
                     also known as p65 (O'Shea and Perkins, 2008)"
                     /label=RelA (p65) AD
                     /translation="PTQAGEGTLSEALLQLQFDDEDLGALLGNSTDPAVFTDLASVDNS
                     EFQQLLNQGIPVAPHTTEPMLMEYPEAITRLVTGAQRPPDPAPAPLGAPGLPNGLLSGD
                     EDFSSIADMDFSALLSQISS"
     protein_bind    complement(2807..2840)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2853..2886
                     /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(2913..3114,3115..3145)
                     /label=P element 3' end
                     /note="P element 3' end"
     primer_bind     complement(3251..3268)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3422..4010)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3502..3521)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4181..4972,4973..5041))
                     /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     4804..4823
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5042..5146)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5309..5330)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(5546..5576,5577..6131)
                     /label=P element 5' end
ORIGIN
        1 ggtaccgggc cccccctcga ggtcgacgaa gttcctattc ttcaaatagt ataggaactt
       61 caagcttctt aaggaattcg agctcgcccg gggatctaat tcaattagag actaattcaa
      121 ttagagctaa ttcaattagg atcagcttat cgatttcgaa ccctcgaccg ccggagtata
      181 aatagaggcg cttcgtctac ggagcgacaa ttcaattcaa acaagcaaag tgaacacgtc
      241 gctaagcgaa agctaagcaa ataaacaagc gcagctgaac aagctaaaca atcgggctag
      301 acctaggcaa aatggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
      361 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
      421 atgccaccta cggcaagctg accctgaagc tcatctgcac caccggcaag ctgcccgtgc
      481 cctggcccac cctcgtgacc accctgggct acggcctgca gtgcttcgcc cgctaccccg
      541 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
      601 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
      661 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
      721 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc accgccgaca
      781 agcagaagaa cggcatcaag gccaacttca agatccgcca caacatcgag gacggcggcg
      841 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
      901 ccgacaacca ctacctgagc taccagtccg ccctgagcaa agaccccaac gagaagcgcg
      961 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     1021 tgtacaagta accatggcta gatcataatc agccatacca catttgtaga ggttttactt
     1081 gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt
     1141 gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     1201 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     1261 gtatcgaatt cgtacgcgcc cggggagccc aagggcacgc cctggcaccc ggcgcgccag
     1321 agggcagagg cagcctgctc acctgcggcg acgtggaaga gaaccctggg cctatgaagg
     1381 ctctcacggc ccgacaacag gaagtttttg atttgatacg ggatcatata tcccaaacgg
     1441 gtatgcctcc gacccgcgca gagatagcac agcgactggg ctttcgatcg cctaacgccg
     1501 cggaggagca cttgaaggca ctggcccgca agggtgtcat tgaaatcgtg tccggtgcga
     1561 gccgcggaat ccggctgttg caggaggagg aagagggcct gccactggtg ggacgcgtgg
     1621 ccgctggcga gccgctgctg gcccagcaac acatagaggg acactatcag gtggacccct
     1681 ccttgtttaa gccaaatgct gatttcctgt tgcgggtgtc gggaatgtcc atgaaggaca
     1741 tcggtattat ggatggtgac ctcctcgccg tccataagac acaggatgtc aggaacggcc
     1801 aggtagtcgt tgccaggata gacgatgagg tcactgtgaa acgtctcaag aagcaaggca
     1861 ataaggtcga gctgctgccg gagaatagcg agttcaagcc gatcgtggtg gatctgcgac
     1921 agcagtcctt tactatcgag ggcttggccg tgggtgtgat ccgcaacgga gattggctgg
     1981 gatccacgcc gatggagttc cagtacctgc ccgatacgga tgaccgtcac cgtatcgaag
     2041 aaaagcggaa gcgaacctat gaaaccttca agtccatcat gaaaaagtcc cccttctcgg
     2101 gccccacgga cccgcgcccc ccgccccgtc gtattgcggt tccttcgcgc agcagtgcca
     2161 gcgtccccaa acccgcaccg cagccctacc cgttcacttc ctcccttagc acgattaact
     2221 atgatgagtt ccccacgatg gtgttcccca gtggacaaat ttcccaggca tcggcactgg
     2281 ctccggcccc accgcaagtc ctcccccagg cgcccgctcc ggcaccggct cccgcaatgg
     2341 tgagtgctct ggcccaggcc cccgctccag tccccgtgct ggcgcctgga cccccacagg
     2401 cagttgcccc tcctgctccg aaaccaacgc aggcgggcga aggaaccctg agcgaggccc
     2461 tcttgcagct tcagttcgat gacgaagact tgggagccct gctgggtaac agcacagacc
     2521 ctgccgtatt caccgatctc gcatccgtgg acaacagcga gtttcagcag ctcttgaatc
     2581 agggaatccc ggtcgcacct cataccacag agcccatgct gatggaatac ccggaggcta
     2641 tcacgcgact ggtgaccggc gcacagcgac cacccgatcc agcccctgcc ccactgggtg
     2701 ccccgggttt gcccaatggc ctcctcagcg gcgatgagga tttctccagc atcgctgata
     2761 tggatttctc cgctttgctg agccagataa gctcctaagc ggccgcataa cttcgtatag
     2821 catacattat acgaagttat accggtggcc gcgaagttcc tattcttcaa atagtatagg
     2881 aacttcgagc tcccggagag atgcagaagc tttgcgtact cgcaaattat taaaaataaa
     2941 actttaaaaa taatttcgtc taattaatat tatgagttaa ttcaaacccc acggacatgc
     3001 taagggttaa tcaacaatca tatcgctgtc tcactcagac tcaatacgac actcagaata
     3061 ctattccttt cactcgcact tattgcaagc atacgttaag tggatgtctc ttgccgacgg
     3121 gaccacctta tgttatttca tcatggtctg gccattctca tcgtgagctt ccgggtgctc
     3181 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     3241 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     3301 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     3361 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     3421 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     3481 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     3541 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     3601 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     3661 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     3721 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     3781 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     3841 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     3901 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     3961 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     4021 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     4081 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     4141 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     4201 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     4261 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     4321 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     4381 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     4441 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     4501 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     4561 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     4621 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     4681 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     4741 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     4801 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     4861 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     4921 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     4981 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     5041 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     5101 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     5161 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     5221 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     5281 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     5341 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     5401 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     5461 taaagggagc ccccgattta gagcttgacg gggaaagccg gccttctccg ccgccgatct
     5521 gccgctggac tacgtgggtc tggcccatga tgaaataaca taaggtggtc ccgtcgatag
     5581 ccgaagctta ccgaagtata cacttaaatt cagtgcacgt ttgcttgttg agaggaaagg
     5641 ttgtgtgcgg acgaattttt ttttgaaaac attaaccctt acgtggaata aaaaaaaatg
     5701 aaatattgca aattttgctg caaagctgtg actggagtaa aattaattca cgtgccgaag
     5761 tgtgctatta agagaaaatt gtgggagcag agccttgggt gcagccttgg tgaaaactcc
     5821 caaatttgtg atacccactt taatgattcg cagtggaagg ctgcacctgc aaaaggtcag
     5881 acatttaaaa ggaggcgact caacgcagat gccgtaccta gtaaagtgat agagcctgaa
     5941 ccagaaaaga taaaagaagg ctataccagt gggagtacac aaacagagta agtttgaata
     6001 gtaaaaaaaa tcatttatgt aaacaataac gtgactgtgc gttaggtcct gttcattgtt
     6061 taatgaaaat aagagcttga gggaaaaaat tcgtactttg gagtacgaaa tgcgtcgttt
     6121 agagcagcag c
//
