LOCUS       sequence-413481-        7705 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Fusion of Ca2+ reporter and endosomal Ca2+-permeable channel.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7705)
  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 7705)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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: TPC1-G-GECO1.2
FEATURES             Location/Qualifiers
     source          1..7705
                     /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             3130..3186
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKTGHAVRAIGRLSS"
     primer_bind     3399..3420
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3928..4371
                     /codon_start=1
                     /product="calcium-binding protein"
                     /label=calmodulin
                     /translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEA
                     ELQDMINEVDADGDGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDGNGYIGAAEL
                     RHVMTNLGEKLTDEEVDEMIRVADIDGDGQVNYEEFVQMMTAK"
     polyA_signal    4493..4614
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4530..4549)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4584..4603
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4621..5076)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4758..4779)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4970..4989
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5103..5207
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5209..5566
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5209..5400
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5232..5252)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5417..5552
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5601..6395
                     /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             5601..6395
                     /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(5655..5674)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6265..6284
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6583..6602)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6627..6674
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7003..7591
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7492..7511
                     /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 atggctgtga gtttggatga cgacgtgccg
      661 ctcatcctga ccttggatga gggtggcagt gccccactgg ctccctccaa cggcctgggc
      721 caagaagagc tacctagcaa aaatggcggc agctatgcca tccacgactc ccaggccccc
      781 agtctcagct ctgggggtga gagttccccc tccagccccg cacacaactg ggagatgaat
      841 taccaagagg cagcaatcta cctccaggaa ggcgagaaca acgacaagtt cttcacccac
      901 cccaaggatg ccaaggcgct ggcggcctac ctctttgcac acaatcacct cttctacctg
      961 atggagctgg ccacggccct gctgctgctg ctgctctccc tgtgcgaggc ccccgccgtc
     1021 cccgcactcc ggcttggcat ctatgtccac gccaccctgg agctgtttgc cctgatggtg
     1081 gtagtgtttg aactctgcat gaagttacgc tggctgggcc tccacacctt catccggcac
     1141 aagcggacca tggtcaagac ctcggtgctg gtggtgcagt ttgtcgaggc catcgtggtg
     1201 ttggtacggc agatgtccca tgtgcgggtg acccgagcac tgcgctgcat tttcctggtg
     1261 gactgtcggt attgcggtgg cgtccggcgc aacctgcggc agatcttcca gtccctgccg
     1321 cccttcatgg acatcctcct gctgctgctg ttcttcatga tcatctttgc catcctcggt
     1381 ttctacttgt tctcccctaa cccttcagac ccctacttca gcaccctgga gaacagcatc
     1441 gtcagtctgt ttgtccttct gaccacagcc aatttcccag atgtgatgat gccctcctac
     1501 tcccggaacc cctggtcctg cgtcttcttc atcgtgtacc tctccatcga gctgtatttc
     1561 atcatgaacc tgcttctggc tgtggtgttc gacaccttca atgacattga gaaacgcaag
     1621 ttcaagtctt tgctactgca caagcgaacc gctatccagc atgcctaccg cctgctcatc
     1681 agccagagga ggcctgccgg catctcctac aggcagtttg aaggcctcat gcgcttctac
     1741 aagccccgga tgagtgccag ggagcgctat cttaccttca aggccctgaa tcagaacaac
     1801 acacccctgc tcagcctaaa ggacttttac gatatctacg aagttgctgc tttgaagtgg
     1861 aaggccaaga aaaacagaga gcactggttt gatgagcttc ccaggacggc gctcctcatc
     1921 ttcaaaggta ttaatatcct tgtgaagtcc aaggccttcc agtatttcat gtacttggtg
     1981 gtggcagtca acggggtctg gatcctcgtg gagacattta tgctgaaagg tgggaacttc
     2041 ttctccaagc acgtgccctg gagttacctc gtctttctaa ctatctatgg ggtggagctg
     2101 ttcctgaagg ttgccggcct gggccctgtg gagtacttgt cttccggatg gaacttgttt
     2161 gacttctccg tgacagtgtt cgccttcctg ggactgctgg cgctggccct caacatggag
     2221 cccttctatt tcatcgtggt cctgcgcccc ctccagctgc tgaggttgtt taagttgaag
     2281 gagcgctacc gcaacgtgct ggacaccatg ttcgagctgc tgccccggat ggccagcctg
     2341 ggcctcaccc tgctcatctt ttactactcc ttcgccatcg tgggcatgga gttcttctgc
     2401 gggatcgtct tccccaactg ctgcaacacg agtacagtgg cagatgccta ccgctggcgc
     2461 aaccacaccg tgggcaacag gaccgtggtg gaggaaggct actattatct caataatttt
     2521 gacaacatcc tcaacagctt tgtgaccctg tttgagctca cagttgtcaa caactggtac
     2581 atcatcatgg aaggcgtcac ctctcagacc tcccactgga gccgcctcta cttcatgacc
     2641 ttttacattg tgaccatggt ggtgatgacg atcattgtcg cctttatcct cgaggccttc
     2701 gtcttccgaa tgaactacag ccgcaagaac caggactcgg aagttgatgg tggcatcacc
     2761 cttgagaagg aaatctccaa agaagagctg gttgccgtcc tggagctcta ccgggaggca
     2821 cggggggcct cctcggatgt caccaggctg ctggagaccc tctcccagat ggagagatac
     2881 cagcaacatt ccatggtgtt tctgggacgg cgatcaagga ccaagagcga cctgagcctg
     2941 aagatgtacc aggaggagat ccaggagtgg tatgaggagc atgccaggga gcaagagcag
     3001 cagcgacaac tcagcagcag tgcagccccc gccgcccagc agcccccagg cagccgccag
     3061 cgctcccaga ccgttaccca gtcgacggta ccgcgggccc gggatccacc ggtcatggtc
     3121 gactcatcac gtcgtaagtg gaataagaca ggtcacgcag tcagagctat aggtcggctg
     3181 agctcactcg agaacgtcta tatcaaggcc gacgagcaga agaacggcat caaggcgtac
     3241 ttcaagatcc gccacaacat cgagggcggc ggcgtgcagc tcgcctacca ctaccagcag
     3301 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcgtgcag
     3361 tccatgcttt cgaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     3421 accgccgccg ggatcactct cggcatggac gagctgtaca agggcggtac cggagggagc
     3481 gaatccatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat ccaggtcgag
     3541 ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggtga gggcgatgcc
     3601 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
     3661 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
     3721 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacatcca ggagcgcacc
     3781 atcttcttca agggcgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
     3841 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
     3901 gggcacaagc tggagtacaa cacgcgtgac caactgactg aagagcagat cgcagaattt
     3961 aaagaggctt tctccctatt tgacaaggac ggggatggga cgataacaac caaggagctg
     4021 gggacggtga tgcggtctct ggggcagaac cccacagaag cagagctgca ggacatgatc
     4081 aatgaagtag atgccgacgg tgacggcaca atcgacttcc ctgagttcct gacaatgatg
     4141 gcaagaaaaa tgaaagacac agacagtgaa gaagaaatta gagaagcgtt ccgtgtgttt
     4201 gataaggacg gcaatggcta catcggcgca gcagagcttc gccacgtgat gacaaacctt
     4261 ggagagaagt taacagatga agaggttgat gaaatgatca gggtagcaga catcgatggg
     4321 gatggtcagg taaactacga agagtttgta caaatgatga cagcgaagta agcggccgcg
     4381 actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct
     4441 cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt
     4501 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4561 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt
     4621 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     4681 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     4741 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     4801 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     4861 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     4921 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     4981 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     5041 cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg
     5101 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     5161 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa
     5221 gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     5281 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     5341 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     5401 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     5461 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt
     5521 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga
     5581 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
     5641 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
     5701 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
     5761 ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg
     5821 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
     5881 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
     5941 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
     6001 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
     6061 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc
     6121 tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
     6181 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
     6241 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg
     6301 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
     6361 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
     6421 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     6481 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     6541 tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac
     6601 aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt
     6661 tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc
     6721 accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc
     6781 aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc
     6841 ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     6901 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     6961 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     7021 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     7081 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     7141 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     7201 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     7261 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     7321 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     7381 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     7441 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     7501 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     7561 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     7621 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     7681 ggataaccgt attaccgcca tgcat
//
