LOCUS       sequence-287405-        5517 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  endoplasmic reticulum-resident FRB for chemically induced 
            dimerization.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5517)
  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 5517)
  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: iRFP-FRB-ER
FEATURES             Location/Qualifiers
     source          1..5517
                     /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"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             613..1560
                     /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"
     CDS             1627..1905
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
     CDS             1639..1908
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="EMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTLKETS
                     FNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISKQ"
     CDS             complement(1919..1936)
                     /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"
     polyA_signal    2305..2426
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2342..2361)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2396..2415
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2433..2888)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2570..2591)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2782..2801
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2915..3019
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3021..3378
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3021..3212
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3044..3064)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3229..3364
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3413..4207
                     /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             3413..4207
                     /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(3467..3486)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4077..4096
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4395..4414)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4439..4486
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4815..5403
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5304..5323
                     /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 gctagcgcta
      601 ccggtcgcca ccatggcgga aggatccgtc gccaggcagc ctgacctctt gacctgcgac
      661 gatgagccga tccatatccc cggtgccatc caaccgcatg gactgctgct cgccctcgcc
      721 gccgacatga cgatcgttgc cggcagcgac aaccttcccg aactcaccgg actggcgatc
      781 ggcgccctga tcggccgctc tgcggccgat gtcttcgact cggagacgca caaccgtctg
      841 acgatcgcct tggccgagcc cggggcggcc gtcggagcac cgatcactgt cggcttcacg
      901 atgcgaaagg acgcaggctt catcggctcc tggcatcgcc atgatcagct catcttcctc
      961 gagctcgagc ctccccagcg ggacgtcgcc gagccgcagg cgttcttccg ccgcaccaac
     1021 agcgccatcc gccgcctgca ggccgccgaa accttggaaa gcgcctgcgc cgccgcggcg
     1081 caagaggtgc ggaagattac cggcttcgat cgggtgatga tctatcgctt cgcctccgac
     1141 ttcagcggcg aagtgatcgc agaggatcgg tgcgccgagg tcgagtcaaa actaggcctg
     1201 cactatcctg cctcaaccgt gccggcgcag gcccgtcggc tctataccat caacccggta
     1261 cggatcattc ccgatatcaa ttatcggccg gtgccggtca ccccagacct caatccggtc
     1321 accgggcggc cgattgatct tagcttcgcc atcctgcgca gcgtctcgcc cgtccatctg
     1381 gaattcatgc gcaacatagg catgcatggc acgatgtcga tctcgatttt gcgcggcgag
     1441 cgactgtggg gattgatcgt ttgccatcac cgaacgccgt actacgtcga tctcgatggc
     1501 cgccaagcct gcgagctagt cgcccaggtt ctggcctggc agatcggcgt gatggaagag
     1561 tccggactca gatctcgagc tcagcttacc atggcttatc cttatgacgt ccctgactat
     1621 gtggccatcc tctggcatga gatgtggcat gaaggcctgg aagaggcatc tcgtttgtac
     1681 tttggggaaa ggaacgtgaa aggcatgttt gaggtgctgg agcccttgca tgctatgatg
     1741 gaacggggcc cccagactct gaaggaaaca tcctttaatc aggcctatgg tcgagattta
     1801 atggaggccc aagagtggtg caggaagtac atgaaatcag ggaatgtcaa ggacctcacc
     1861 caagcctggg acctctatta tcatgtgttc cgacgaatct caaagcaggg atccggtgct
     1921 ggtgctggtg ctggtgctat cctgaattcc cgggtcttcc tggctttgcc tgttatcatg
     1981 gttgttgcct tttcaatgtg cattatctgt ttgcttatgg ctggtgacac ttggacagaa
     2041 acactggcct atgtgctctt ctggggagtt gcaagcattg gaacattttt tatcattctt
     2101 tacaatggca aagattttgt cgatgctccc agactggtcc agaaagaaaa gatagactga
     2161 ggtaccgcgg gcccgggatc caccggatct agataactga tcataatcag ccataccaca
     2221 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     2281 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     2341 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     2401 ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta agcgttaata ttttgttaaa
     2461 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     2521 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     2581 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     2641 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     2701 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     2761 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     2821 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     2881 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     2941 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     3001 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
     3061 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3121 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3181 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     3241 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     3301 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     3361 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga
     3421 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     3481 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     3541 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
     3601 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     3661 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     3721 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     3781 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     3841 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     3901 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
     3961 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     4021 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     4081 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     4141 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     4201 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     4261 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     4321 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct
     4381 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
     4441 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt
     4501 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
     4561 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
     4621 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt
     4681 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     4741 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     4801 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     4861 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     4921 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     4981 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     5041 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     5101 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     5161 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     5221 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     5281 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     5341 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     5401 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     5461 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat
//
