LOCUS       sequence-360219-        5787 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PtdIns(3,4,5)P3 biosensor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5787)
  AUTHORS   Walpole GFW, Pacheco J, Chauhan N, Clark J, Anderson KE, Abbas YM, 
            Brabant-Kirwan D, Montano-Rendon F, Liu Z, Zhu H, Brumell JH, 
            Deiters A, Stephens LR, Hawkins PT, Hammond GRV, Grinstein S, Fairn 
            GD
  TITLE     Kinase-independent synthesis of 3-phosphorylated phosphoinositides 
            by a phosphotransferase.
  JOURNAL   Nat Cell Biol. 2022 Apr 28. pii: 10.1038/s41556-022-00895-y. doi: 
            10.1038/s41556-022-00895-y.
  PUBMED    35484249
REFERENCE   2  (bases 1 to 5787)
  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: "Nat Cell 
            Biol. 2022 Apr 28. pii: 10.1038/s41556-022-00895-y. doi: 
            10.1038/s41556-022-00895-y."
COMMENT     Sequence Label: NES-mCherry-PH-BTKx2
FEATURES             Location/Qualifiers
     source          1..5787
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        73..376
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        377..580
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     531..551
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      664..673
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             670..1377
                     /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(815..833)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1083..1102
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2587..2708
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2624..2643)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2678..2697
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2715..3170)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2852..2873)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3064..3083
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3197..3301
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3303..3660
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3303..3494
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3326..3346)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3511..3646
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3695..4489
                     /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             3695..4489
                     /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(3749..3768)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4359..4378
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4677..4696)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4721..4768
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5097..5685
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5586..5605
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 accgccatgc attagttatt aatagtaatc aattacgggg tcattagttc atagcccata
       61 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      121 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      181 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      241 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      301 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      361 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      421 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      481 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      541 cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat
      601 ccgctagcca ccatggctct gcagaaaaag ttggaagagc ttgagctgga tgaggcaccg
      661 gtcgccacca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg
      721 cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag
      781 ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc
      841 cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac
      901 gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag
      961 tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc
     1021 ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac
     1081 ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc
     1141 gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac
     1201 tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc
     1261 tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa
     1321 cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtcc
     1381 ggactcagat ctcgagctca agcttcgaat tctgccgcag tgattctgga gagcatcttt
     1441 ctgaagcgat cccaacagaa aaagaaaaca tcacctctaa acttcaagaa gcgcctgttt
     1501 ctcttgaccg tgcacaaact ctcctactat gagtatgact ttgaacgtgg gagaagaggc
     1561 agtaagaagg gttcaataga tgttgagaag atcacttgtg ttgaaacagt ggttcctgaa
     1621 aaaaatcctc ctccagaaag acagattccg agaagaggtg aagagtccag tgaaatggag
     1681 caaatttcaa tcattgaaag gttcccttat cccttccagg ttgtatatga tgaagggcct
     1741 ctctacgtct tctccccaac tgaagaacta aggaagcggt ggattcacca gctcaaaaac
     1801 gtaatccggt acaacagtga tctggttcag aaatatcacc cttgcttctg gatcgatggg
     1861 cagtatctct gctgctctca gacagccaaa aatgctatgg gctgccaaat tttggagaac
     1921 aggaatggaa gcgggggttc cggcggagcc gcagtgattc tggagagcat ctttctgaag
     1981 cgatcccaac agaaaaagaa aacatcacct ctaaacttca agaagcgcct gtttctcttg
     2041 accgtgcaca aactctccta ctatgagtat gactttgaac gtgggagaag aggcagtaag
     2101 aagggttcaa tagatgttga gaagatcact tgtgttgaaa cagtggttcc tgaaaaaaat
     2161 cctcctccag aaagacagat tccgagaaga ggtgaagagt ccagtgaaat ggagcaaatt
     2221 tcaatcattg aaaggttccc ttatcccttc caggttgtat atgatgaagg gcctctctac
     2281 gtcttctccc caactgaaga actaaggaag cggtggattc accagctcaa aaacgtaatc
     2341 cggtacaaca gtgatctggt tcagaaatat cacccttgct tctggatcga tgggcagtat
     2401 ctctgctgct ctcagacagc caaaaatgct atgggctgcc aaattttgga gaactgagga
     2461 tccaccggat ctagataact gatcataatc agccatacca catttgtaga ggttttactt
     2521 gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt
     2581 gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     2641 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     2701 gtatcttaac gcgtaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg
     2761 ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa
     2821 agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa
     2881 gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg
     2941 tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa
     3001 ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa
     3061 ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct
     3121 gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac
     3181 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     3241 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     3301 tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     3361 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     3421 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     3481 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     3541 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg
     3601 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     3661 gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
     3721 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
     3781 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
     3841 caagaccgac ctgtccggtg ccctgaatga actgcaagac gaggcagcgc ggctatcgtg
     3901 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
     3961 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
     4021 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
     4081 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
     4141 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
     4201 actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg
     4261 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     4321 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     4381 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     4441 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     4501 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     4561 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     4621 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag gctaactgaa
     4681 acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat
     4741 aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac
     4801 tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc
     4861 ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca
     4921 ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa acttcatttt
     4981 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     5041 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     5101 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     5161 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     5221 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     5281 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     5341 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     5401 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     5461 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     5521 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     5581 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     5641 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     5701 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     5761 tcccctgatt ctgtggataa ccgtatt
//
