LOCUS       sequence-399146-        6357 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 6357)
  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 6357)
  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-PIP4K2AD273K
FEATURES             Location/Qualifiers
     source          1..6357
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..378
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        21..212
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(44..64)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      229..364
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             413..1207
                     /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             413..1207
                     /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(467..486)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1077..1096
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1395..1414)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1439..1486
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1815..2403
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2304..2323
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2578..2881
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2882..3085
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3036..3056
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      3124..3133
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3130..3837
                     /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(3275..3293)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3543..3562
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             3898..4218
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     polyA_signal    5662..5783
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5699..5718)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5753..5772
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5790..6245)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5927..5948)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6139..6158
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(6272..6357,1..19)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
       61 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
      121 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
      181 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
      241 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
      301 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
      361 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga
      421 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
      481 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
      541 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
      601 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
      661 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
      721 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
      781 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
      841 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
      901 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
      961 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     1021 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     1081 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     1141 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     1201 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     1261 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     1321 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct
     1381 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
     1441 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt
     1501 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
     1561 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
     1621 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt
     1681 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     1741 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     1801 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     1861 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     1921 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     1981 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     2041 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     2101 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     2161 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     2221 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     2281 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     2341 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     2401 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     2461 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcattag
     2521 ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt
     2581 tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac
     2641 gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg
     2701 ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag
     2761 tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat
     2821 gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat
     2881 ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt
     2941 tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga
     3001 ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg
     3061 gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct agcgctaccg
     3121 gtcgccacca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg
     3181 cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag
     3241 ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc
     3301 cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac
     3361 gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag
     3421 tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc
     3481 ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac
     3541 ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc
     3601 gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac
     3661 tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc
     3721 tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa
     3781 cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtcc
     3841 ggactcagat ctcgaagcgc ggccgcggga gcaggaggag cagctcgagc ggcgctggga
     3901 gtgcaggtgg aaaccatctc cccaggagac gggcgcacct tccccaagcg cggccagacc
     3961 tgcgtggtgc actacaccgg gatgcttgaa gatggaaaga aatttgattc ctcccgggac
     4021 agaaacaagc cctttaagtt tatgctaggc aagcaggagg tgatccgagg ctgggaagaa
     4081 ggggttgccc agatgagtgt gggtcagaga gccaaactga ctatatctcc agattatgcc
     4141 tatggtgcca ctgggcaccc aggcatcatc ccaccacatg ccactctcgt cttcgatgtg
     4201 gagcttctaa aactggaaag tgctggtggt agtgctggtg gtagtgctgg tggtagtgct
     4261 ggtggtagtg ctggtggtcc gcgagctcaa gcttcgaatt ctgcagtcga catggcgacc
     4321 cccggcaacc tagggtcctc cgtcctggcg agcaagacca agaccaagaa gaagcacttc
     4381 gtagcgcaga aagtgaagct gtttcgggcc agcgacccgc tgctcagcgt cctcatgtgg
     4441 ggggtaaacc actcgatcaa tgaactgagc catgttcaaa tccctgttat gttgatgcca
     4501 gatgacttca aagcctattc aaaaataaag gtggacaatc acctttttaa caaagaaaac
     4561 atgccgagcc atttcaagtt taaggaatac tgcccgatgg tcttccgtaa cctgcgggag
     4621 aggtttggaa ttgatgatca agatttccag aattccctga ccaggagcgc acccctcccc
     4681 aacgactccc aggcccgcag tggagctcgt tttcacactt cctacgacaa aagatacatc
     4741 atcaagacta ttaccagtga agacgtggcc gaaatgcaca acatcctgaa gaaataccac
     4801 cagtacatag tggaatgtca tgggatcacc cttcttcccc agttcttggg catgtaccgg
     4861 cttaatgttg atggagttga aatatatgtg atagttacaa gaaatgtatt cagccaccgt
     4921 ttgtctgtgt ataggaaata cgacttaaag ggctctacag tggctagaga agctagtgac
     4981 aaagaaaagg ccaaagaact gccaactctg aaagataatg atttcattaa tgagggccaa
     5041 aagatttata ttgatgacaa cagcaagaag gtcttcctgg aaaaactaaa aaaggatgtt
     5101 gagtttctgg cccagctgaa gctcatgaaa tacagtctgc tggtgggaat tcatgatgtg
     5161 gagagagccg aacaggagga agtggagtgt gaggagaacg atggggagga ggagggcgag
     5221 agcgatggca cccacccggt gggaaccccc ccagatagcc ccgggaatac actgaacagc
     5281 tcaccacccc tggctcccgg ggagttcgat ccgaacatcg acgtctatgg aattaagtgc
     5341 catgaaaact cgcctaggaa ggaggtgtac ttcatggcaa ttattgacat ccttactcat
     5401 tatgatgcaa aaaagaaagc tgcccatgct gcaaaaactg ttaaacatgg cgctggcgcg
     5461 gagatctcca ccgtgaaccc agaacagtat tcaaagcgct ttttggactt tattggccac
     5521 atcttgacgt aaggatccac cggatctaga taactgatca taatcagcca taccacattt
     5581 gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa
     5641 atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc
     5701 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     5761 tccaaactca tcaatgtatc ttaacgcgta aattgtaagc gttaatattt tgttaaaatt
     5821 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     5881 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     5941 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     6001 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     6061 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     6121 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     6181 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     6241 cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     6301 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaata
//
