LOCUS       sequence-399280-        6413 bp    DNA     circular SYN 15-APR-2025
DEFINITION  expresses recruitable fluorescent protein-tagged enzyme.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6413)
  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 6413)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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-PIP4K2CD280K
FEATURES             Location/Qualifiers
     source          1..6413
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             69..863
                     /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             69..863
                     /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(123..142)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     733..752
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1051..1070)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1095..1142
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1471..2059
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1960..1979
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2234..2537
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2538..2741
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2692..2712
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2780..2789
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2786..3493
                     /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(2931..2949)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3199..3218
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             3554..3874
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     polyA_signal    5374..5495
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5411..5430)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5465..5484
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5502..5957)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5639..5660)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5851..5870
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5984..6088
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(6090..6413,1..34)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6090..6281
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6113..6133)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(6298..6413,1..20)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat
       61 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
      121 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
      181 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
      241 atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
      301 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
      361 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
      421 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
      481 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
      541 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca
      601 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
      661 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
      721 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
      781 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
      841 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
      901 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
      961 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     1021 gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg
     1081 aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt
     1141 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
     1201 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
     1261 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc
     1321 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     1381 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     1441 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     1501 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     1561 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     1621 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     1681 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     1741 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     1801 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     1861 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     1921 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     1981 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     2041 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     2101 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     2161 taccgccatg cattagttat taatagtaat caattacggg gtcattagtt catagcccat
     2221 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
     2281 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
     2341 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
     2401 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
     2461 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
     2521 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
     2581 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
     2641 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
     2701 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg aaccgtcaga
     2761 tccgctagcg ctaccggtcg ccaccatggt gagcaagggc gaggaggata acatggccat
     2821 catcaaggag ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt
     2881 cgagatcgag ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa
     2941 ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta
     3001 cggctccaag gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt
     3061 ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt
     3121 gacccaggac tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac
     3181 caacttcccc tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc
     3241 cgagcggatg taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct
     3301 gaaggacggc ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt
     3361 gcagctgccc ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga
     3421 ctacaccatc gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga
     3481 cgagctgtac aagtccggac tcagatctcg aagcgcggcc gcgggagcag gaggagcagc
     3541 tcgagcggcg ctgggagtgc aggtggaaac catctcccca ggagacgggc gcaccttccc
     3601 caagcgcggc cagacctgcg tggtgcacta caccgggatg cttgaagatg gaaagaaatt
     3661 tgattcctcc cgggacagaa acaagccctt taagtttatg ctaggcaagc aggaggtgat
     3721 ccgaggctgg gaagaagggg ttgcccagat gagtgtgggt cagagagcca aactgactat
     3781 atctccagat tatgcctatg gtgccactgg gcacccaggc atcatcccac cacatgccac
     3841 tctcgtcttc gatgtggagc ttctaaaact ggaaagtgct ggtggtagtg ctggtggtag
     3901 tgctggtggt agtgctggtg gtagtgctgg tggtccgcga gctcaagctt cgaattctgc
     3961 agtcgacggt accgcgatgg cgtcctcctc ggtcccacca gccacggtat cggcggcgac
     4021 agcaggcccc ggcccaggtt tcggcttcgc ctccaagacc aagaagaagc atttcgtgca
     4081 gcagaaggtg aaggtgttcc gggcggccga cccgctggtg ggtgtgttcc tgtggggcgt
     4141 agcccactcg atcaatgagc tcagccaggt gcctcccccg gtgatgctgc tgccagatga
     4201 ctttaaggcc agctccaaga tcaaggtcaa caatcacctt ttccacaggg aaaatctgcc
     4261 cagtcatttc aagttcaagg agtattgtcc ccaggtcttc aggaacctcc gtgatcgatt
     4321 tggcattgat gaccaagatt acttggtgtc ccttacccga aaccccccca gcgaaagtga
     4381 aggcagtgat ggtcgcttcc ttatctccta cgatcggact ctggtcatca aagaagtatc
     4441 cagtgaggac attgctgaca tgcatagcaa cctctccaac tatcaccagt acattgtgaa
     4501 gtgccatggc aacacgcttc tgccccagtt cctggggatg taccgagtca gtgtggacaa
     4561 cgaagacagc tacatgcttg tgatgcgcaa tatgtttagc caccgtcttc ctgtgcacag
     4621 gaagtatgac ctcaagggtt ccctagtgtc ccgggaagcc agcgataagg aaaaggttaa
     4681 agaattgccc acccttaagg atatggactt tctcaacaag aaccagaaag tatatattgg
     4741 tgaagaggag aagaaaatat ttctggagaa gctgaagaga gatgtggagt ttctagtgca
     4801 gctgaagatc atgaaataca gccttctgct aggcatccac gacatcattc ggggctctga
     4861 accagaggag gaagcgcccg tgcgggagga tgagtcagag gtggatgggg actgcagcct
     4921 gactggacct cctgctctgg tgggctccta tggcacctcc ccagagggta tcggaggcta
     4981 catccattcc catcggcccc tgggcccagg agagtttgag tccttcattg atgtctatgc
     5041 catccggagt gctgaaggag ccccccagaa ggaggtctac ttcatgggcc tcattgatat
     5101 ccttacacag tatgatgcca agaagaaagc agctcatgca gccaaaactg tcaagcatgg
     5161 ggctggggca gagatctcta ctgtccatcc ggagcagtat gctaagcgat tcctggattt
     5221 tattaccaac atctttgcct aacgggatcc accggatcta gataactgat cataatcagc
     5281 cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac
     5341 ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt
     5401 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     5461 agttgtggtt tgtccaaact catcaatgta tcttaacgcg taaattgtaa gcgttaatat
     5521 tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
     5581 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc
     5641 agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac
     5701 cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc
     5761 gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg
     5821 gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag
     5881 ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc
     5941 gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     6001 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     6061 gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct gtggaatgtg
     6121 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     6181 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     6241 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     6301 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     6361 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtg
//
