LOCUS       sequence-399474-        6431 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses recruitable fluorescent protein-tagged enzyme.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6431)
  AUTHORS   Wills RC, Doyle CP, Zewe JP, Pacheco J, Hansen SD, Hammond GRV
  TITLE     A novel homeostatic mechanism tunes PI(4,5)P2-dependent signaling at
            the plasma membrane.
  JOURNAL   J Cell Sci. 2023 Aug 3:jcs.261494. doi: 10.1242/jcs.261494.
  PUBMED    37534432
REFERENCE   2  (bases 1 to 6431)
  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 Sci.
            2023 Aug 3:jcs.261494. doi: 10.1242/jcs.261494."
COMMENT     Sequence Label: iRFP713-FRB-Mss4-Kina
FEATURES             Location/Qualifiers
     source          1..6431
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             102..896
                     /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             102..896
                     /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(156..175)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     766..785
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1084..1103)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1128..1175
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1504..2092
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1993..2012
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2267..2570
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2571..2774
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2725..2745
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2813..2822
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2819..3766
                     /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             3815..4093
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
     polyA_signal    5425..5546
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5462..5481)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5516..5535
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5553..6008)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5690..5711)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5902..5921
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6035..6139
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(6141..6431,1..67)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6141..6332
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6164..6184)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(6349..6431,1..53)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
       61 ttgcaaagat cgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat
      121 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
      181 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
      241 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc
      301 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
      361 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc
      421 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
      481 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
      541 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
      601 cagccgaact gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga
      661 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
      721 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
      781 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
      841 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg
      901 gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga
      961 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     1021 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccta gggggaggct
     1081 aactgaaaca cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag
     1141 acagaataaa acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg
     1201 ctggcactct gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc
     1261 cttttcccca ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg
     1321 ggcggcaggc cctgccatag cctcaggtta ctcatatata ctttagattg atttaaaact
     1381 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     1441 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     1501 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     1561 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     1621 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     1681 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     1741 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     1801 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     1861 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     1921 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     1981 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     2041 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     2101 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     2161 tgcgttatcc cctgattctg tggataaccg tattaccgcc atgcattagt tattaatagt
     2221 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
     2281 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
     2341 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
     2401 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
     2461 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
     2521 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
     2581 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
     2641 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
     2701 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
     2761 atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg tcgccaccat
     2821 ggcggaagga tccgtcgcca ggcagcctga cctcttgacc tgcgacgatg agccgatcca
     2881 tatccccggt gccatccaac cgcatggact gctgctcgcc ctcgccgccg acatgacgat
     2941 cgttgccggc agcgacaacc ttcccgaact caccggactg gcgatcggcg ccctgatcgg
     3001 ccgctctgcg gccgatgtct tcgactcgga gacgcacaac cgtctgacga tcgccttggc
     3061 cgagcccggg gcggccgtcg gagcaccgat cactgtcggc ttcacgatgc gaaaggacgc
     3121 aggcttcatc ggctcctggc atcgccatga tcagctcatc ttcctcgagc tcgagcctcc
     3181 ccagcgggac gtcgccgagc cgcaggcgtt cttccgccgc accaacagcg ccatccgccg
     3241 cctgcaggcc gccgaaacct tggaaagcgc ctgcgccgcc gcggcgcaag aggtgcggaa
     3301 gattaccggc ttcgatcggg tgatgatcta tcgcttcgcc tccgacttca gcggcgaagt
     3361 gatcgcagag gatcggtgcg ccgaggtcga gtcaaaacta ggcctgcact atcctgcctc
     3421 aaccgtgccg gcgcaggccc gtcggctcta taccatcaac ccggtacgga tcattcccga
     3481 tatcaattat cggccggtgc cggtcacccc agacctcaat ccggtcaccg ggcggccgat
     3541 tgatcttagc ttcgccatcc tgcgcagcgt ctcgcccgtc catctggaat tcatgcgcaa
     3601 cataggcatg cacggcacga tgtcgatctc gattttgcgc ggcgagcgac tgtggggatt
     3661 gatcgtttgc catcaccgaa cgccgtacta cgtcgatctc gatggccgcc aagcctgcga
     3721 gctagtcgcc caggttctgg cctggcagat cggcgtgatg gaagagtccg gactcagatc
     3781 tcgagctgga ggtgctggtg ctatcctatc tagaatcctc tggcatgaga tgtggcatga
     3841 aggcctggaa gaggcatctc gtttgtactt tggggaaagg aacgtgaaag gcatgtttga
     3901 ggtgctggag cccttgcatg ctatgatgga acggggcccc cagactctga aggaaacatc
     3961 ctttaatcag gcctatggtc gagatttaat ggaggcccaa gagtggtgca ggaagtacat
     4021 gaaatcaggg aatgtcaagg acctcaccca agcctgggac ctctattatc atgtgttccg
     4081 acgaatctca aagggtggta gtgctggtgg tagtgctcaa gcttcgaatt ctgcagtcga
     4141 cggtaccgcg aacaaggtct cagagggtca cgtgaatttc atcattgcct acaacatgct
     4201 cactggcata agagttgccg tcagtagatg ttctgggatc atgaagccac tcaccccagc
     4261 agattttagg tttaccaaaa aactggcctt tgattatcac ggcaatgagc ttactccgtc
     4321 atcccagtat gcatttaagt tcaaggatta ttgtcccgag gttttccggg agcttagagc
     4381 actgtttggc ctcgatcctg ccgattatct ggtgtctctc acatctaaat acatcctctc
     4441 agaactgaat agcccaggaa aatctgggtc atttttttat tatagtcgcg attacaagta
     4501 tattattaag actattcacc actctgaaca catccacctc cggaaacaca tccaagaata
     4561 ctataaccat gtccgcgata acccgaatac tcttatttgc caattttatg gcctgcatcg
     4621 cgtcaagatg ccaatctcct tccagaacaa aatcaaacat cgaaaaatat atttcctggt
     4681 gatgaacaac ctgttccccc cacatcttga tattcacatc acctatgatc tcaagggatc
     4741 cacatggggc aggttcacca atcttgacaa agagcgcttg gctaaagatc gcagctaccg
     4801 cccagtgatg aaagacctta actggctgga ggagggtcaa aagattaagt tcgggccact
     4861 gaagaaaaag acattcctta cccagcttaa gaaggatgta gagcttctgg ccaaactgaa
     4921 cacaatggat tacagcctgc ttattggcat ccacgatatc aataaagcca aagaggacga
     4981 cttgcaattg gcggacactg ctagcattga agaacagcct caaacgcagg gccctattag
     5041 gacaggcaca ggcacagttg tacggcattt cttccgcgag ttcgagggcg gcataagagc
     5101 tagcgaccag tttaacaacg acgtcgacct tatttattat gtggggataa tcgactttct
     5161 gacaaactac agtgtcatga aaaaactcga gactttttgg cgctcacttc gacacgacac
     5221 taaattggtc agcgctatac ctccacggga ttatgctaat cgcttttacg agtttatcga
     5281 ggattccgtc taacgggatc caccggatct agataactga tcataatcag ccataccaca
     5341 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     5401 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     5461 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     5521 ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta agcgttaata ttttgttaaa
     5581 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     5641 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     5701 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     5761 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     5821 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     5881 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     5941 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     6001 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     6061 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     6121 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
     6181 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     6241 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     6301 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     6361 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     6421 gaggccgagg c
//
