LOCUS       sequence-413484-        7513 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fusion of Ca2+ reporter and pore-dead lysosomal Ca2+-permeable 
            channel.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7513)
  AUTHORS   Davis LC, Morgan AJ, Galione A
  TITLE     NAADP-regulated two-pore channels drive phagocytosis through 
            endo-lysosomal Ca(2+) nanodomains, calcineurin and dynamin.
  JOURNAL   EMBO J. 2020 Jul 15;39(14):e104058. doi: 10.15252/embj.2019104058. 
            Epub 2020 Jun 8.
  PUBMED    32510172
REFERENCE   2  (bases 1 to 7513)
  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: "EMBO J."; 
            date: "2020-07-15"; volume: "39(14):e104058. doi"; pages: " 
            10.15252/embj.2019104058. Epub 2020 Jun 8"
COMMENT     Sequence Label: TPC2-D276K-G-GECO1.2
FEATURES             Location/Qualifiers
     source          1..7513
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             2938..2994
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKTGHAVRAIGRLSS"
     primer_bind     3207..3228
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3736..4179
                     /codon_start=1
                     /product="calcium-binding protein"
                     /label=calmodulin
                     /translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEA
                     ELQDMINEVDADGDGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDGNGYIGAAEL
                     RHVMTNLGEKLTDEEVDEMIRVADIDGDGQVNYEEFVQMMTAK"
     polyA_signal    4301..4422
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4338..4357)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4392..4411
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4429..4884)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4566..4587)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4778..4797
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4911..5015
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5017..5374
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5017..5208
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5040..5060)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5225..5360
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5409..6203
                     /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             5409..6203
                     /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(5463..5482)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6073..6092
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6391..6410)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6435..6482
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6811..7399
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7300..7319
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttacc atggcggaac cccaggcgga gtcggagccc
      661 ctgctgggcg gggcccgcgg cggtggcggc gactggccgg cggggctgac cacttaccgc
      721 agcatccaag tcggccctgg tgccgcggcc aggtgggacc tctgcattga tcaggctgtg
      781 gtcttcatcg aagatgctat tcagtaccgc tccatcaacc accgggtgga tgccagctcg
      841 atgtggcttt accgacggta ttactcgaac gtatgccaac ggactttgag cttcaccatc
      901 ttcttgatcc tgtttttggc ttttatcgag accccatcct cactcaccag cacggcggac
      961 gtgcgctacc gcgctgcccc ctgggagccg ccctgcggcc tgaccgagag tgtcgaggtg
     1021 ctctgcctgc tggtctttgc ggccgacctc tctgtgaagg gttacctgtt cgggtgggcc
     1081 catttccaga aaaacctttg gctgctgggc tacctcgtgg tgctggtggt gtctctggtg
     1141 gactggaccg tgtccctgag tctcgtgtgt catgagcccc tgcggatccg ccggcttctc
     1201 cgtcccttct tcctgctgca gaactcctct atgatgaaga agaccttgaa atgcatccgc
     1261 tggtcgctgc cggaaatggc cagcgtcggg ctgctgctgg ccatccacct gtgcctcttc
     1321 accatgttcg gaatgctgct gttcgctggt gggaagcagg atgatgggca ggacagggag
     1381 aggctgacct acttccagaa cctgcctgag tctctgactt ccctcctggt gctgctgacc
     1441 acggccaaca accccaaagt gatgattcct gcgtattcca agaaccgggc ctatgccatc
     1501 ttcttcatag tcttcactgt gataggaagc ctgtttctga tgaacctgct gacagccatc
     1561 atctacagtc agttccgggg ctacctgatg aaatctctcc agacctcgct gtttcggagg
     1621 cggctgggaa cccgggctgc ctttgaagtc ctatcctcca tggtggggga gggaggagcc
     1681 ttccctcagg cagttggggt gaagccccag aacttgctgc aggtgcttca gaaggtccag
     1741 ctggacagct cccacaaaca ggccatgatg gagaaggtgc gttcctacgg cagtgttctg
     1801 ctgtcagctg aggagtttca gaagctcttc aacgagcttg acagaagtgt ggttaaagag
     1861 cacccgccga ggcccgagta ccagtctccg tttctgcaga gcgcccagtt cctcttcggc
     1921 cactactact ttgactacct ggggaacctc atcgccctgg caaacctggt gtccatttgc
     1981 gtgttcctgg tgctggatgc agatgtgctg cctgctgagc gtgatgactt catcctgggg
     2041 attctcaact gcgtcttcat tgtgtactac ctgttggaga tgctgctcaa ggtctttgcc
     2101 ctgggcctgc gagggtacct gtcctacccc agcaacgtgt ttgacgggct cctcaccgtt
     2161 gtcctgctgg ttttggagat ctcaactctg gctgtgtacc gattgccaca cccaggctgg
     2221 aggccggaga tggtgggcct gctgtcgctg tgggacatga cccgcatgct gaacatgctc
     2281 atcgtgttcc gcttcctgcg tatcatcccc agcatgaagc cgatggccgt ggtggccagt
     2341 accgtcctgg gcctggtgca gaacatgcgt gcttttggcg ggatcctggt ggtggtctac
     2401 tacgtatttg ccatcattgg gatcaacttg tttagaggcg tcattgtggc tcttcctgga
     2461 aacagcagcc tggcccctgc caatggctcg gcgccctgtg ggagcttcga gcagctggag
     2521 tactgggcca acaacttcga tgactttgcg gctgccctgg tcactctgtg gaacttgatg
     2581 gtggtgaaca actggcaggt gtttctggat gcatatcggc gctactcagg cccgtggtcc
     2641 aagatctatt ttgtattgtg gtggctggtg tcgtctgtca tctgggtcaa cctgtttctg
     2701 gccctgattc tggagaactt ccttcacaag tgggaccccc gcagccacct gcagcccctt
     2761 gctgggaccc cagaggccac ctaccagatg actgtggagc tcctgttcag ggatattctg
     2821 gaggagcccg aggaggatga gctcacagag aggctgagcc agcacccgca cctgtggctg
     2881 tgcaggcagt cgacggtacc gcgggcccgg gatccaccgg tcatggtcga ctcatcacgt
     2941 cgtaagtgga ataagacagg tcacgcagtc agagctatag gtcggctgag ctcactcgag
     3001 aacgtctata tcaaggccga cgagcagaag aacggcatca aggcgtactt caagatccgc
     3061 cacaacatcg agggcggcgg cgtgcagctc gcctaccact accagcagaa cacccccatc
     3121 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcgtgcagtc catgctttcg
     3181 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     3241 atcactctcg gcatggacga gctgtacaag ggcggtaccg gagggagcga atccatggtg
     3301 agcaagggcg aggagctgtt caccggggtg gtgcccatcc aggtcgagct ggacggcgac
     3361 gtaaacggcc acaagttcag cgtgtccggc gagggtgagg gcgatgccac ctacggcaag
     3421 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     3481 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     3541 gacttcttca agtccgccat gcccgaaggc tacatccagg agcgcaccat cttcttcaag
     3601 ggcgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     3661 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     3721 gagtacaaca cgcgtgacca actgactgaa gagcagatcg cagaatttaa agaggctttc
     3781 tccctatttg acaaggacgg ggatgggacg ataacaacca aggagctggg gacggtgatg
     3841 cggtctctgg ggcagaaccc cacagaagca gagctgcagg acatgatcaa tgaagtagat
     3901 gccgacggtg acggcacaat cgacttccct gagttcctga caatgatggc aagaaaaatg
     3961 aaagacacag acagtgaaga agaaattaga gaagcgttcc gtgtgtttga taaggacggc
     4021 aatggctaca tcggcgcagc agagcttcgc cacgtgatga caaaccttgg agagaagtta
     4081 acagatgaag aggttgatga aatgatcagg gtagcagaca tcgatgggga tggtcaggta
     4141 aactacgaag agtttgtaca aatgatgaca gcgaagtaag cggccgcgac tctagatcat
     4201 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
     4261 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
     4321 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     4381 gcattctagt tgtggtttgt ccaaactcat caatgtatct taaggcgtaa attgtaagcg
     4441 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
     4501 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
     4561 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
     4621 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
     4681 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
     4741 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
     4801 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
     4861 ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     4921 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     4981 gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg
     5041 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     5101 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     5161 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     5221 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     5281 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
     5341 aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat
     5401 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     5461 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     5521 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     5581 atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     5641 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     5701 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     5761 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     5821 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     5881 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca
     5941 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     6001 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     6061 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     6121 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     6181 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     6241 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     6301 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     6361 gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg
     6421 aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt
     6481 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
     6541 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
     6601 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc
     6661 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     6721 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     6781 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     6841 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     6901 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     6961 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     7021 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     7081 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     7141 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     7201 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     7261 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     7321 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     7381 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     7441 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     7501 taccgccatg cat
//
