LOCUS       sequence-399136-        5978 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses fluorescent protein-tagged enzyme.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5978)
  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 5978)
  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: TagBFP2-PIP4K2C
FEATURES             Location/Qualifiers
     source          1..5978
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        63..420
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        63..254
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(86..106)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      271..406
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             455..1249
                     /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             455..1249
                     /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(509..528)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1119..1138
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1437..1456)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1481..1528
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1857..2445
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2346..2365
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2620..2923
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2924..3127
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3078..3098
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      3166..3175
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3172..3873
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIK
                     VVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMAL
                     KLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARY
                     CDLPSKLGHKLN"
     polyA_signal    5325..5446
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5362..5381)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5416..5435
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5453..5908)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5590..5611)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5802..5821
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(5935..5978,1..61)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
       61 tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
      121 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
      181 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
      241 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
      301 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg
      361 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
      421 gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
      481 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
      541 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
      601 caagaccgac ctgtccggtg ccctgaatga actgcaagac gaggcagcgc ggctatcgtg
      661 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
      721 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
      781 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
      841 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
      901 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
      961 actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg
     1021 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     1081 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     1141 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     1201 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     1261 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     1321 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     1381 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag gctaactgaa
     1441 acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat
     1501 aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac
     1561 tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc
     1621 ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca
     1681 ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa acttcatttt
     1741 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     1801 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     1861 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     1921 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     1981 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     2041 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     2101 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     2161 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     2221 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     2281 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     2341 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     2401 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     2461 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     2521 tcccctgatt ctgtggataa ccgtattacc gccatgcatt agttattaat agtaatcaat
     2581 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     2641 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     2701 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     2761 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     2821 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     2881 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
     2941 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
     3001 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     3061 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     3121 cagagctggt ttagtgaacc gtcagatccg ctagcgctac cggtcgccac catggtgagc
     3181 gagctgatta aggagaacat gcacatgaag ctgtacatgg agggcaccgt ggacaaccat
     3241 cacttcaagt gcacatccga gggcgaaggc aagccctacg agggcaccca gaccatgaga
     3301 atcaaggtgg tcgagggcgg ccctctcccc ttcgccttcg acatcctggc tactagcttc
     3361 ctctacggca gcaagacctt catcaaccac acccagggca tccccgactt cttcaagcag
     3421 tccttccctg agggcttcac atgggagaga gtcaccacat acgaagacgg gggcgtgctg
     3481 accgctaccc aggacaccag cctccaggac ggctgcctca tctacaacgt caagatcaga
     3541 ggggtgaact tcacatccaa cggccctgtg atgcagaaga aaacactcgg ctgggaggcc
     3601 ttcaccgaga cgctgtaccc cgctgacggc ggcctggaag gcagaaacga catggccctg
     3661 aagctcgtgg gcgggagcca tctgatcgca aacgccaaga ccacatatag atccaagaaa
     3721 cccgctaaga acctcaagat gcctggcgtc tactatgtgg actacagact ggaaagaatc
     3781 aaggaggcca acaacgagac ctacgtcgag cagcacgagg tggcagtggc cagatactgc
     3841 gacctcccta gcaaactggg gcacaagctt aattccggac tcagatctcg agctcaagct
     3901 tcgaattctg cagtcgacgg taccgcgatg gcgtcctcct cggtcccacc agccacggta
     3961 tcggcggcga cagcaggccc cggcccaggt ttcggcttcg cctccaagac caagaagaag
     4021 catttcgtgc agcagaaggt gaaggtgttc cgggcggccg acccgctggt gggtgtgttc
     4081 ctgtggggcg tagcccactc gatcaatgag ctcagccagg tgcctccccc ggtgatgctg
     4141 ctgccagatg actttaaggc cagctccaag atcaaggtca acaatcacct tttccacagg
     4201 gaaaatctgc ccagtcattt caagttcaag gagtattgtc cccaggtctt caggaacctc
     4261 cgtgatcgat ttggcattga tgaccaagat tacttggtgt cccttacccg aaaccccccc
     4321 agcgaaagtg aaggcagtga tggtcgcttc cttatctcct acgatcggac tctggtcatc
     4381 aaagaagtat ccagtgagga cattgctgac atgcatagca acctctccaa ctatcaccag
     4441 tacattgtga agtgccatgg caacacgctt ctgccccagt tcctggggat gtaccgagtc
     4501 agtgtggaca acgaagacag ctacatgctt gtgatgcgca atatgtttag ccaccgtctt
     4561 cctgtgcaca ggaagtatga cctcaagggt tccctagtgt cccgggaagc cagcgataag
     4621 gaaaaggtta aagaattgcc cacccttaag gatatggact ttctcaacaa gaaccagaaa
     4681 gtatatattg gtgaagagga gaagaaaata tttctggaga agctgaagag agatgtggag
     4741 tttctagtgc agctgaagat catggactac agccttctgc taggcatcca cgacatcatt
     4801 cggggctctg aaccagagga ggaagcgccc gtgcgggagg atgagtcaga ggtggatggg
     4861 gactgcagcc tgactggacc tcctgctctg gtgggctcct atggcacctc cccagagggt
     4921 atcggaggct acatccattc ccatcggccc ctgggcccag gagagtttga gtccttcatt
     4981 gatgtctatg ccatccggag tgctgaagga gccccccaga aggaggtcta cttcatgggc
     5041 ctcattgata tccttacaca gtatgatgcc aagaagaaag cagctcatgc agccaaaact
     5101 gtcaagcatg gggctggggc agagatctct actgtccatc cggagcagta tgctaagcga
     5161 ttcctggatt ttattaccaa catctttgcc taacgggatc caccggatct agataactga
     5221 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
     5281 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     5341 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     5401 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta
     5461 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     5521 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     5581 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     5641 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     5701 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     5761 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     5821 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     5881 gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga
     5941 acccctattt gtttattttt ctaaatacat tcaaatat
//
