LOCUS       sequence-399756-        6261 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 6261)
  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 6261)
  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: mCherry-FKBP-PIP4K2A-Kina
FEATURES             Location/Qualifiers
     source          1..6261
                     /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..1320
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(758..776)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1026..1045
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1381..1701
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     polyA_signal    3049..3170
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3086..3105)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3140..3159
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3177..3632)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3314..3335)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3526..3545
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3659..3763
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3765..4122
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3765..3956
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3788..3808)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3973..4108
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4157..4951
                     /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             4157..4951
                     /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(4211..4230)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4821..4840
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5139..5158)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5183..5230
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5559..6147
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6048..6067
                     /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 ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc
      661 atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc
      721 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt
      781 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc
      841 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc
      901 aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc
      961 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc
     1021 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac
     1081 cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc
     1141 cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc
     1201 gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg
     1261 gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag
     1321 tccggactca gatctcgaag cgcggccgcg ggagcaggag gagcagctcg agcggcgctg
     1381 ggagtgcagg tggaaaccat ctccccagga gacgggcgca ccttccccaa gcgcggccag
     1441 acctgcgtgg tgcactacac cgggatgctt gaagatggaa agaaatttga ttcctcccgg
     1501 gacagaaaca agccctttaa gtttatgcta ggcaagcagg aggtgatccg aggctgggaa
     1561 gaaggggttg cccagatgag tgtgggtcag agagccaaac tgactatatc tccagattat
     1621 gcctatggtg ccactgggca cccaggcatc atcccaccac atgccactct cgtcttcgat
     1681 gtggagcttc taaaactgga aagtgctggt ggtagtgctg gtggtagtgc tggtggtagt
     1741 gctggtggta gtgctggtgg tccgcgagct caagcttcga attctgcagt cgacgccagc
     1801 gacccgctgc tcagcgtcct catgtggggg gtaaaccact cgatcaatga actgagccat
     1861 gttcaaatcc ctgttatgtt gatgccagat gacttcaaag cctattcaaa aataaaggtg
     1921 gacaatcacc tttttaacaa agaaaacatg ccgagccatt tcaagtttaa ggaatactgc
     1981 ccgatggtct tccgtaacct gcgggagagg tttggaattg atgatcaaga tttccagaat
     2041 tccctgacca ggagcgcacc cctccccaac gactcccagg cccgcagtgg agctcgtttt
     2101 cacacttcct acgacaaaag atacatcatc aagactatta ccagtgaaga cgtggccgaa
     2161 atgcacaaca tcctgaagaa ataccaccag tacatagtgg aatgtcatgg gatcaccctt
     2221 cttccccagt tcttgggcat gtaccggctt aatgttgatg gagttgaaat atatgtgata
     2281 gttacaagaa atgtattcag ccaccgtttg tctgtgtata ggaaatacga cttaaagggc
     2341 tctacagtgg ctagagaagc tagtgacaaa gaaaaggcca aagaactgcc aactctgaaa
     2401 gataatgatt tcattaatga gggccaaaag atttatattg atgacaacag caagaaggtc
     2461 ttcctggaaa aactaaaaaa ggatgttgag tttctggccc agctgaagct catggactac
     2521 agtctgctgg tgggaattca tgatgtggag agagccgaac aggaggaagt ggagtgtgag
     2581 gagaacgatg gggaggagga gggcgagagc gatggcaccc acccggtggg aaccccccca
     2641 gatagccccg ggaatacact gaacagctca ccacccctgg ctcccgggga gttcgatccg
     2701 aacatcgacg tctatggaat taagtgccat gaaaactcgc ctaggaagga ggtgtacttc
     2761 atggcaatta ttgacatcct tactcattat gatgcaaaaa agaaagctgc ccatgctgca
     2821 aaaactgtta aacatggcgc tggcgcggag atctccaccg tgaacccaga acagtattca
     2881 aagcgctttt tggactttat tggccacatc ttgacgtaag gatccaccgg atctagataa
     2941 ctgatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
     3001 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
     3061 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     3121 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat
     3181 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     3241 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     3301 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     3361 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     3421 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     3481 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     3541 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     3601 cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg
     3661 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     3721 ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac
     3781 cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     3841 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     3901 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3961 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     4021 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     4081 aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg
     4141 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     4201 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     4261 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     4321 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     4381 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     4441 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     4501 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     4561 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     4621 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     4681 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     4741 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     4801 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     4861 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     4921 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     4981 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     5041 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     5101 atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata
     5161 ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg
     5221 tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg
     5281 agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt
     5341 tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca
     5401 ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     5461 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     5521 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     5581 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     5641 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     5701 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     5761 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     5821 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     5881 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     5941 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     6001 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     6061 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     6121 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     6181 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     6241 aaccgtatta ccgccatgca t
//
