LOCUS       sequence-399220-        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- PIP4K2CD280K
FEATURES             Location/Qualifiers
     source          1..5978
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        57..414
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        57..248
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(80..100)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      265..400
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             449..1243
                     /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             449..1243
                     /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(503..522)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1113..1132
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1431..1450)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1475..1522
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1851..2439
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2340..2359
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2614..2917
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2918..3121
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3072..3092
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      3160..3169
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3166..3867
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIK
                     VVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMAL
                     KLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARY
                     CDLPSKLGHKLN"
     polyA_signal    5319..5440
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5356..5375)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5410..5429
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5447..5902)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5584..5605)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5796..5815
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(5929..5978,1..55)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga
       61 ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
      121 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
      181 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
      241 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
      301 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
      361 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga
      421 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
      481 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
      541 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
      601 cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc
      661 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
      721 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
      781 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
      841 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
      901 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
      961 cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     1021 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     1081 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     1141 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     1201 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     1261 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     1321 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     1381 cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg
     1441 aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg
     1501 cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc
     1561 gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc
     1621 caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct
     1681 gccatagcct caggttactc atatatactt tagattgatt taaaacttca tttttaattt
     1741 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     1801 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     1861 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     1921 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     1981 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     2041 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     2101 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     2161 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     2221 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     2281 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     2341 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     2401 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     2461 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     2521 gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat caattacggg
     2581 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     2641 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
     2701 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
     2761 ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga
     2821 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg
     2881 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat
     2941 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
     3001 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc
     3061 cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
     3121 tggtttagtg aaccgtcaga tccgctagcg ctaccggtcg ccaccatggt gagcgagctg
     3181 attaaggaga acatgcacat gaagctgtac atggagggca ccgtggacaa ccatcacttc
     3241 aagtgcacat ccgagggcga aggcaagccc tacgagggca cccagaccat gagaatcaag
     3301 gtggtcgagg gcggccctct ccccttcgcc ttcgacatcc tggctactag cttcctctac
     3361 ggcagcaaga ccttcatcaa ccacacccag ggcatccccg acttcttcaa gcagtccttc
     3421 cctgagggct tcacatggga gagagtcacc acatacgaag acgggggcgt gctgaccgct
     3481 acccaggaca ccagcctcca ggacggctgc ctcatctaca acgtcaagat cagaggggtg
     3541 aacttcacat ccaacggccc tgtgatgcag aagaaaacac tcggctggga ggccttcacc
     3601 gagacgctgt accccgctga cggcggcctg gaaggcagaa acgacatggc cctgaagctc
     3661 gtgggcggga gccatctgat cgcaaacgcc aagaccacat atagatccaa gaaacccgct
     3721 aagaacctca agatgcctgg cgtctactat gtggactaca gactggaaag aatcaaggag
     3781 gccaacaacg agacctacgt cgagcagcac gaggtggcag tggccagata ctgcgacctc
     3841 cctagcaaac tggggcacaa gcttaattcc ggactcagat ctcgagctca agcttcgaat
     3901 tctgcagtcg acggtaccgc gatggcgtcc tcctcggtcc caccagccac ggtatcggcg
     3961 gcgacagcag gccccggccc aggtttcggc ttcgcctcca agaccaagaa gaagcatttc
     4021 gtgcagcaga aggtgaaggt gttccgggcg gccgacccgc tggtgggtgt gttcctgtgg
     4081 ggcgtagccc actcgatcaa tgagctcagc caggtgcctc ccccggtgat gctgctgcca
     4141 gatgacttta aggccagctc caagatcaag gtcaacaatc accttttcca cagggaaaat
     4201 ctgcccagtc atttcaagtt caaggagtat tgtccccagg tcttcaggaa cctccgtgat
     4261 cgatttggca ttgatgacca agattacttg gtgtccctta cccgaaaccc ccccagcgaa
     4321 agtgaaggca gtgatggtcg cttccttatc tcctacgatc ggactctggt catcaaagaa
     4381 gtatccagtg aggacattgc tgacatgcat agcaacctct ccaactatca ccagtacatt
     4441 gtgaagtgcc atggcaacac gcttctgccc cagttcctgg ggatgtaccg agtcagtgtg
     4501 gacaacgaag acagctacat gcttgtgatg cgcaatatgt ttagccaccg tcttcctgtg
     4561 cacaggaagt atgacctcaa gggttcccta gtgtcccggg aagccagcga taaggaaaag
     4621 gttaaagaat tgcccaccct taaggatatg gactttctca acaagaacca gaaagtatat
     4681 attggtgaag aggagaagaa aatatttctg gagaagctga agagagatgt ggagtttcta
     4741 gtgcagctga agatcatgaa atacagcctt ctgctaggca tccacgacat cattcggggc
     4801 tctgaaccag aggaggaagc gcccgtgcgg gaggatgagt cagaggtgga tggggactgc
     4861 agcctgactg gacctcctgc tctggtgggc tcctatggca cctccccaga gggtatcgga
     4921 ggctacatcc attcccatcg gcccctgggc ccaggagagt ttgagtcctt cattgatgtc
     4981 tatgccatcc ggagtgctga aggagccccc cagaaggagg tctacttcat gggcctcatt
     5041 gatatcctta cacagtatga tgccaagaag aaagcagctc atgcagccaa aactgtcaag
     5101 catggggctg gggcagagat ctctactgtc catccggagc agtatgctaa gcgattcctg
     5161 gattttatta ccaacatctt tgcctaacgg gatccaccgg atctagataa ctgatcataa
     5221 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc
     5281 tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata
     5341 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     5401 attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat tgtaagcgtt
     5461 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     5521 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     5581 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     5641 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     5701 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     5761 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     5821 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     5881 aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     5941 atttgtttat ttttctaaat acattcaaat atgtatcc
//
