LOCUS       sequence-399525-        5952 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 5952)
  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 5952)
  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-PIP4K2B
FEATURES             Location/Qualifiers
     source          1..5952
                     /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..3520
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIK
                     VVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMAL
                     KLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARY
                     CDLPSKLGHKLN"
     polyA_signal    4946..5067
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4983..5002)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5037..5056
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5074..5529)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5211..5232)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5423..5442
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5556..5660
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(5662..5952,1..67)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5662..5853
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5685..5705)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(5870..5952,1..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 ggtgagcgag ctgattaagg agaacatgca catgaagctg tacatggagg gcaccgtgga
     2881 caaccatcac ttcaagtgca catccgaggg cgaaggcaag ccctacgagg gcacccagac
     2941 catgagaatc aaggtggtcg agggcggccc tctccccttc gccttcgaca tcctggctac
     3001 tagcttcctc tacggcagca agaccttcat caaccacacc cagggcatcc ccgacttctt
     3061 caagcagtcc ttccctgagg gcttcacatg ggagagagtc accacatacg aagacggggg
     3121 cgtgctgacc gctacccagg acaccagcct ccaggacggc tgcctcatct acaacgtcaa
     3181 gatcagaggg gtgaacttca catccaacgg ccctgtgatg cagaagaaaa cactcggctg
     3241 ggaggccttc accgagacgc tgtaccccgc tgacggcggc ctggaaggca gaaacgacat
     3301 ggccctgaag ctcgtgggcg ggagccatct gatcgcaaac gccaagacca catatagatc
     3361 caagaaaccc gctaagaacc tcaagatgcc tggcgtctac tatgtggact acagactgga
     3421 aagaatcaag gaggccaaca acgagaccta cgtcgagcag cacgaggtgg cagtggccag
     3481 atactgcgac ctccctagca aactggggca caagcttaat tccggactca gatctcgagc
     3541 tcaagcttcg aattctgcag tcgacatgtc gtccaactgc accagcacca cggcggtggc
     3601 ggtggcgccg ctcagcgcca gcaagaccaa gaccaagaag aagcatttcg tgtgccagaa
     3661 agtgaagcta ttccgggcca gcgagccgat cctcagcgtc ctgatgtggg gggtgaacca
     3721 cacgatcaat gagctgagca atgttcctgt tcctgtcatg ctaatgccag atgacttcaa
     3781 agcctacagc aagatcaagg tggacaatca tctcttcaat aaggagaacc tgcccagccg
     3841 ctttaagttt aaggagtatt gccccatggt gttccgaaac cttcgggaga ggtttggaat
     3901 tgatgatcag gattaccaga attcagtgac gcgcagcgcc cccatcaaca gtgacagcca
     3961 gggtcggtgt ggcacgcgtt tcctcaccac ctacgaccgg cgctttgtca tcaagactgt
     4021 gtccagcgag gacgtggcgg agatgcacaa catcttaaag aaataccacc agtttatagt
     4081 ggagtgtcat ggcaacacgc ttttgccaca gttcctgggc atgtaccgcc tgaccgtgga
     4141 tggtgtggaa acctacatgg tggttaccag gaacgtgttc agccatcggc tcactgtgca
     4201 tcgcaagtat gacctcaagg gttctacggt tgccagagaa gcgagcgaca aggagaaggc
     4261 caaggacttg ccaacattca aagacaatga cttcctcaat gaagggcaga agctgcatgt
     4321 gggagaggag agtaaaaaga acttcctgga gaaactgaag cgggacgttg agttcttggc
     4381 acagctgaag atcatggact acagcctgct ggtgggcatc cacgacgtgg accgggcaga
     4441 gcaggaggag atggaggtgg aggagcgggc agaggacgag gagtgtgaga atgatggggt
     4501 gggtggcaac ctactctgct cctatggcac acctccggac agccctggca acctcctcag
     4561 ctttcctcgg ttctttggtc ctggggaatt cgacccctct gttgacgtct atgccatgaa
     4621 aagccatgaa agttccccca agaaggaggt gtatttcatg gccatcattg atatcctcac
     4681 gccatacgat acaaagaaga aagctgcaca tgctgccaaa acggtgaaac acggggcagg
     4741 ggccgagatc tcgactgtga accctgagca gtactccaaa cgcttcaacg agtttatgtc
     4801 caacatcctg acgtagggat ccaccggatc tagataactg atcataatca gccataccac
     4861 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
     4921 taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata
     4981 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     5041 tttgtccaaa ctcatcaatg tatcttaacg cgtaaattgt aagcgttaat attttgttaa
     5101 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     5161 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     5221 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     5281 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     5341 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     5401 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     5461 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     5521 agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     5581 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     5641 aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt
     5701 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     5761 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     5821 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     5881 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     5941 agaggccgag gc
//
