LOCUS       sequence-407125-        6349 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FRB expression in Schneider cells for imaging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6349)
  AUTHORS   Salgania HK, Metz J, Jeske M
  TITLE     ReLo is a simple and rapid colocalization assay to identify and 
            characterize direct protein-protein interactions.
  JOURNAL   Nat Commun. 2024 Apr 3;15(1):2875. doi: 10.1038/s41467-024-47233-4.
  PUBMED    38570497
REFERENCE   2  (bases 1 to 6349)
  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: "Nat 
            Commun."; date: "2024-04-3"; volume: "15(1):2875. doi"; pages: " 
            10.1038/s41467-024-47233-4"
COMMENT     Sequence Label: pAc5.1-mEGFP-human FRB (HK168)
FEATURES             Location/Qualifiers
     source          1..6349
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        8..2517
                     /label=Ac5 promoter
                     /note="Drosophila melanogaster actin 5C promoter"
     CDS             join(2537..2539,2540..2542,2543..3253)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2582..2603)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2843..2862)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3190..3211
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3278..3556
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
     CDS             3607..3648
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             3658..3675
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3698..3715)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3748..3882
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3772..3791)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3826..3845
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(4483..5071)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4563..4582)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5242..6033,6034..6102))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5865..5884
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6103..6207)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6275..6293
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 aaaaaatcat gaatggcatc aactctgaat caaatctttg cagatgcacc tacttctcat
       61 ttccactgtc acatcatttt tccagatctc gctgcctgtt atgtggccca caaaccaaga
      121 cacgttttat ggccattaaa gctggctgat cgtcgccaaa caccaaatac ataatgaata
      181 tgtacacatt cgagaaagaa gcgatcaaag aagcgtcttc gggcggagta ggagaatgcg
      241 gaggagaagg agaacgagct gatctagtat ctctccacaa tccaatgcca actgaccaac
      301 tggccatatt cggagcaatt tgaagccaat ttccatcgcc tggcgatcgc tccattcttg
      361 gctatatgtt tttcaccgtt acccggggcc attttcaaag actcgtcggc aagataagat
      421 tgtgtcactc gctgtctctc ttcatttgtc gaagaatgct gaggaatttc gcgatgacgt
      481 cggcgagtat tttgaagaat gagaataatt tgtatttata cgaaaatcag ttagtggaat
      541 tttctacaaa aacatgttat ctatagataa ttttgttgca aaatatgttg actatgacaa
      601 agattgtatg tatatacctt taatgtattc taattttctt atgtatttat aatggcaatg
      661 atgatactga tgatatttta agatgatgcc agaccaaaag gcttgaattt ctgcgtcttt
      721 tgccgaacgc agtgcatgtg caattgttgt tttttggaat attcaatttt cggactgtcc
      781 gctttgattt cagtttcttg gcttattcaa aaagcaaagt aaagccaaaa aagcgagatg
      841 gcaataccaa atgcggcaaa acggtagtgg aaggaaaggg gtgcggggca gcggaaggaa
      901 gggtggggcg gggcgtggcg gggtctgtgg ctgggcgcga cgtcaccgac gttggagcca
      961 ctcctttgac catgtgtgcg tgtgtgtatt attcgtgtct cgccactcgc cggttgtttt
     1021 tttcttttta tgctgcgctc tctctagcgc catctcgctt acgcatgctc aacgcaccgc
     1081 atgttgccgt ttccttttat gcgtcatttt ggctcgaaat aggcaattat ttaaacaaag
     1141 attagtcaac gaaaacgcta aaataaataa gtctacaata tggttactta ttgccatgtg
     1201 tgtgcagcca acgatagcaa caaaagcaac aacacaggtg gctttccctc tttcactttt
     1261 tgtttgcaag ccgcgtgcga gcaagacggc acgaccggca aacgcaatta cgctgacaaa
     1321 gagcagacga agttttggcg aaaaacatca aggcgcctga tacgaatgca tttgcaataa
     1381 caattgcgat atttaatatt gtttatgaag ctgtttgact tcaaaacaca caaaaaaaaa
     1441 aataaaacaa attatttgaa agagaattag gaatcggacg cttatcgtta gggtaacaac
     1501 aagaaatgct tactgagtca cagcctctgg aaaactgccg caagccagag agagagagaa
     1561 aaagagggag agcagcttag accgcatgtg cttgtgtgtg aggcgtctct ctcttcgtct
     1621 ctgttgcgca aacgcataga ctgcactgaa aaaatcgatt acctattttt tatgaatgaa
     1681 tatttgcact attactattc aaaactatta agatagcaat cacattcaat agccaaatac
     1741 tataccacct gagcgatgca acgaaatgat caatttgagc aaaaatgctg catatttagg
     1801 acggcatcat tatagaaatg cttcttgctg tgtacttttc tctcgtctgg cagctgtttc
     1861 gccgttattg ttaaaaccgg cttaagttag gtgtgttttc tacgactagt gaatgcccta
     1921 ctagaagatg tgtgttgcac aaaatgtccc tggaataacc aatttgaagt gcagatagca
     1981 gtaaacgtaa gctaatatga atattattta actgtaatgt tttaatatcg ctggacatta
     2041 ctaataaacc cactataaac acatgtacat atgtatgttt tggcatacaa tgagtagttg
     2101 gggaaaaaat gtgtaaaagc accgtgacca tcacagcata aagataacca gctgaagtat
     2161 cgaatatgag taacccccaa attgaatcac atgccgcaac tgataggacc catggaagta
     2221 cactcttcat ggcgatatac aagacacaca caagcacgaa cacccagttg cggaggaaat
     2281 tctccgtaaa tgaaaaccca atcggcgaac aattcatacc catatatggt aaaagttttg
     2341 aacgcgactt gagagcggag agcattgcgg ctgataaggt tttagcgcta agcgggcttt
     2401 ataaaacggg ctgcgggacc agttttcata tcactaccgt ttgagttctt gtgctgtgtg
     2461 gatactcctc ccgacacaaa gccgctccat cagccagcag tcgtctaatc cagagacccc
     2521 ggatcggggt accatcatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
     2581 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga
     2641 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     2701 cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta
     2761 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
     2821 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
     2881 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     2941 caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc
     3001 cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg
     3061 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
     3121 gctgcccgac aaccactacc tgagcaccca gtccaagctg agcaaagacc ccaacgagaa
     3181 gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga
     3241 cgagctgtac aagccaagct tgaattctgc agatatgatc ctctggcatg agatgtggca
     3301 tgaaggcctg gaagaggcat ctcgtttgta ctttggggaa aggaacgtga aaggcatgtt
     3361 tgaggtgctg gagcccttgc atgctatgat ggaacggggc ccccagactc tgaaggaaac
     3421 atcctttaat caggcctatg gtcgagattt aatggaggcc caagagtggt gcaggaagta
     3481 catgaaatca gggaatgtca aggacctcac ccaagcctgg gacctctatt atcatgtgtt
     3541 ccgacgaatc tcaaagtaat ccagcacagt ggcggccgct cgagtctaga gggcccgcgg
     3601 ttcgaaggta agcctatccc taaccctctc ctcggtctcg attctacgcg taccggtcat
     3661 catcaccatc accattgagt ttaaacccgc tgatcagcct cgactgtgcc ttctaaggcc
     3721 tgagctcgct gatcagcctc gatcgaggat ccagacatga taagatacat tgatgagttt
     3781 ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct
     3841 attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt
     3901 cattttatgt ttcaggttca gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc
     3961 tacaaatgtg gtatggctga ttatgatcag tcgaccgatg cccttgagag ccttcaaccc
     4021 agtcagctcc ttccggtggg cgcggggcat gactatcgtc gccgcactta tgactgtctt
     4081 ctttatcatg caactcgtag gacaggtgcc ggcagcgctc tgggtcattt tcggcgagga
     4141 ccgctttcgc tggagcgcga cgatgatcgg cctgtcgctt gcggtattcg gaatcttgca
     4201 cgccctcgct caagccttcg tcactggtcc cgccaccaaa cgtttcggcg agaagcaggc
     4261 cattatcgcc ggcatggcgg ccgacgcgct gggctacgtc ttgctggcgt tcgcgacgcg
     4321 aggctggatg gccttcccca ttatgattct tctcgcttcc ggcggcatcg ggatgcccgc
     4381 gttgcaggcc atgctgtcca ggcaggtaga tgacgaccat cagggacagc ttcaaggcca
     4441 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     4501 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     4561 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4621 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     4681 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     4741 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     4801 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     4861 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     4921 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     4981 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     5041 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5101 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5161 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     5221 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     5281 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     5341 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     5401 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     5461 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     5521 gccagttaat agtttgcgca acgttgttgc cattgctgca ggcatcgtgg tgtcacgctc
     5581 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     5641 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     5701 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     5761 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     5821 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaaca cgggataata ccgcgccaca
     5881 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     5941 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     6001 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     6061 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     6121 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     6181 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta
     6241 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
     6301 tcttcaagaa ttaattctat attctaaaaa cacaaatgat acttctaaa
//
