LOCUS       sequence-287401-        5359 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mitochondrial-resident FRB for chemically induced dimerization.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5359)
  AUTHORS   Zewe JP, Miller AM, Sangappa S, Wills RC, Goulden BD, Hammond GRV
  TITLE     Probing the subcellular distribution of phosphatidylinositol reveals
            a surprising lack at the plasma membrane.
  JOURNAL   J Cell Biol. 2020 Mar 2;219(3). pii: 133808. doi: 
            10.1083/jcb.201906127.
  PUBMED    32211893
REFERENCE   2  (bases 1 to 5359)
  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: "J Cell 
            Biol."; date: "2020-03-2"; volume: "219(3). pii: 133808. doi"; 
            pages: " 10.1083/jcb.201906127"
COMMENT     Sequence Label: Akap1(31)-FRB-iRFP
FEATURES             Location/Qualifiers
     source          1..5359
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             complement(718..735)
                     /codon_start=1
                     /product="HQHQHQ metal affinity tag"
                     /label=HQ tag
                     /note="The HQ tag binds to nickel and other metal ions much
                     like the 6xHis tag, but may allow elution at lower 
                     concentrations of imidazole (Godat et al., 2008)."
                     /translation="HQHQHQ"
     CDS             750..1028
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
     CDS             1035..1061
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      1068..1077
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1074..2024
                     /codon_start=1
                     /product="phytochrome-based near-infrared fluorescent 
                     protein, originally termed iRFP (Filonov et al., 2011)"
                     /label=iRFP713
                     /note="name was changed to iRFP713 (Shcherbakova and 
                     Verkhusha, 2013)"
                     /translation="MAEGSVARQPDLLTCDDEPIHIPGAIQPHGLLLALAADMTIVAGS
                     DNLPELTGLAIGALIGRSAADVFDSETHNRLTIALAEPGAAVGAPITVGFTMRKDAGFI
                     GSWHRHDQLIFLELEPPQRDVAEPQAFFRRTNSAIRRLQAAETLESACAAAAQEVRKIT
                     GFDRVMIYRFASDFSGEVIAEDRCAEVESKLGLHYPASTVPAQARRLYTINPVRIIPDI
                     NYRPVPVTPDLNPVTGRPIDLSFAILRSVSPVHLEFMRNIGMHGTMSISILRGERLWGL
                     IVCHHRTPYYVDLDGRQACELVAQVLAWQIGVMEE"
     polyA_signal    2147..2268
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2184..2203)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2238..2257
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2275..2730)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2412..2433)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2624..2643
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2757..2861
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2863..3220
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2863..3054
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2886..2906)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3071..3206
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3255..4049
                     /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             3255..4049
                     /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(3309..3328)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3919..3938
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4237..4256)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4281..4328
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4657..5245
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5146..5165
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcacca
      601 tggcaatcca gttgcgttcg ctcttcccct tggcgttgcc cggactgctg gccctccttg
      661 gctggtggtg gtttttctct cgtaaaaagg atcccaccag atctgcgaat tcaggtgctg
      721 gtgctggtgc tggtgctatc ctatctagaa tcctctggca tgagatgtgg catgaaggcc
      781 tggaagaggc atctcgtttg tactttgggg aaaggaacgt gaaaggcatg tttgaggtgc
      841 tggagccctt gcatgctatg atggaacggg gcccccagac tctgaaggaa acatccttta
      901 atcaggccta tggtcgagat ttaatggagg cccaagagtg gtgcaggaag tacatgaaat
      961 cagggaatgt caaggacctc acccaagcct gggacctcta ttatcatgtg ttccgacgaa
     1021 tctcaaagac tagttatccg tacgacgtac cagactacgc accggtcgcc accatggcgg
     1081 aaggatccgt cgccaggcag cctgacctct tgacctgcga cgatgagccg atccatatcc
     1141 ccggtgccat ccaaccgcat ggactgctgc tcgccctcgc cgccgacatg acgatcgttg
     1201 ccggcagcga caaccttccc gaactcaccg gactggcgat cggcgccctg atcggccgct
     1261 ctgcggccga tgtcttcgac tcggagacgc acaaccgtct gacgatcgcc ttggccgagc
     1321 ccggggcggc cgtcggagca ccgatcactg tcggcttcac gatgcgaaag gacgcaggct
     1381 tcatcggctc ctggcatcgc catgatcagc tcatcttcct cgagctcgag cctccccagc
     1441 gggacgtcgc cgagccgcag gcgttcttcc gccgcaccaa cagcgccatc cgccgcctgc
     1501 aggccgccga aaccttggaa agcgcctgcg ccgccgcggc gcaagaggtg cggaagatta
     1561 ccggcttcga tcgggtgatg atctatcgct tcgcctccga cttcagcggc gaagtgatcg
     1621 cagaggatcg gtgcgccgag gtcgagtcaa aactaggcct gcactatcct gcctcaaccg
     1681 tgccggcgca ggcccgtcgg ctctatacca tcaacccggt acggatcatt cccgatatca
     1741 attatcggcc ggtgccggtc accccagacc tcaatccggt caccgggcgg ccgattgatc
     1801 ttagcttcgc catcctgcgc agcgtctcgc ccgtccatct ggaattcatg cgcaacatag
     1861 gcatgcacgg cacgatgtcg atctcgattt tgcgcggcga gcgactgtgg ggattgatcg
     1921 tttgccatca ccgaacgccg tactacgtcg atctcgatgg ccgccaagcc tgcgagctag
     1981 tcgcccaggt tctggcctgg cagatcggcg tgatggaaga gtgaagcggc cgcgactcta
     2041 gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca
     2101 cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc
     2161 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     2221 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaag gcgtaaattg
     2281 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     2341 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     2401 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     2461 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     2521 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     2581 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     2641 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     2701 ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg
     2761 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     2821 aaccctgata aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca
     2881 gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     2941 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     3001 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     3061 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     3121 actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa
     3181 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat
     3241 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     3301 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     3361 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     3421 ccctgaatga actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     3481 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     3541 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     3601 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     3661 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     3721 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     3781 cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     3841 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     3901 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     3961 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     4021 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca
     4081 agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt
     4141 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
     4201 gctggagttc ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc
     4261 ggaaggaacc cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc
     4321 gtttgttcat aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag
     4381 accccattgg ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc
     4441 gggtgaaggc ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg
     4501 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     4561 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     4621 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     4681 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     4741 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     4801 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     4861 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     4921 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     4981 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     5041 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     5101 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     5161 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     5221 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     5281 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     5341 ccgtattacc gccatgcat
//
