LOCUS       sequence-315201-        5054 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express cpFRB2 as part of a circularly permuted rapamycin-inducible 
            dimerization system (cpRAPID); visualized by mCherry2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5054)
  AUTHORS   Lee YT, He L, Zhou Y
  TITLE     Expanding the Chemogenetic Toolbox by Circular Permutation.
  JOURNAL   J Mol Biol. 2020 May 1;432(10):3127-3136. doi: 
            10.1016/j.jmb.2020.03.033. Epub 2020 Apr 8.
  PUBMED    32277990
REFERENCE   2  (bases 1 to 5054)
  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.jmb.2020.03.033"; journalName: "J Mol Biol"; date: 
            "2020-05-1- 1"; volume: "432"; issue: "10"; pages: "3127-3136"
COMMENT     Sequence Label: mCh-cpFRB2
FEATURES             Location/Qualifiers
     source          1..5054
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      547..556
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             553..1260
                     /codon_start=1
                     /product="derivative of the monomeric red fluorescent 
                     protein mCherry"
                     /label=mCherry2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFNWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQCRTMGWEASTERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVDIKLDILSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(698..716)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      1291..1300
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1782..1903
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1819..1838)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1873..1892
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1910..2365)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2047..2068)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2259..2278
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2392..2496
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2498..2855
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2498..2689
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2521..2541)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2706..2841
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2890..3684
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2890..3684
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2944..2963)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3554..3573
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3872..3891)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3916..3963
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4292..4880
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4781..4800
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggtcgcca ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc
      601 atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc
      661 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt
      721 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc
      781 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc
      841 aattgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc
      901 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc
      961 gacggccccg taatgcagtg tcgtaccatg ggctgggagg cctccactga gcggatgtac
     1021 cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc
     1081 cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc
     1141 gcctacaacg tcgacatcaa gttggacatc ctttcccaca acgaggacta caccatcgtg
     1201 gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag
     1261 tccggactca gatctcgagc tcaagcttca gccaccatgg gtaatgttaa agatttgact
     1321 caagcgtggg acctgtatta tcacgtcttt cgccggatta gtaaacaagg tggtagcgga
     1381 ggatcaatgg aacttataag agtacctatc ttgtggcatg agatgtggca cgagggattg
     1441 gaagaagcgt cacgccttta tttcggcgaa cgcaacgtta agggtatgtt tgaggtcctg
     1501 gagccactgc acgccatgat ggagcgcgga ccccagactc tgaaggagac tagttttaat
     1561 caggcgtatg gacgggatct catggaggcg caggagtggt gccggaagta catgaaatca
     1621 cgaattctgc agtcgacggt accgcgggcc cgggatccac cggatctaga taactgatca
     1681 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     1741 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     1801 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     1861 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaacgcgta aattgtaagc
     1921 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     1981 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     2041 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     2101 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     2161 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     2221 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     2281 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     2341 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     2401 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     2461 tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt
     2521 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     2581 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     2641 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     2701 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     2761 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     2821 gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga
     2881 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     2941 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     3001 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     3061 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     3121 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     3181 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     3241 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     3301 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     3361 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     3421 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     3481 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     3541 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     3601 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     3661 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     3721 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     3781 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     3841 agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag
     3901 gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg
     3961 ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc
     4021 attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg
     4081 aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact
     4141 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     4201 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     4261 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     4321 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     4381 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     4441 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     4501 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     4561 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     4621 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     4681 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     4741 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     4801 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     4861 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     4921 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     4981 ttaccgccat gcattagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
     5041 tatatggagt tccg
//
