LOCUS       sequence-267156-        6991 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses tagged TPC2 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6991)
  AUTHORS   Davis LC, Morgan AJ, Chen JL, Snead CM, Bloor-Young D, Shenderov E, 
            Stanton-Humphreys MN, Conway SJ, Churchill GC, Parrington J, 
            Cerundolo V, Galione A
  TITLE     NAADP activates two-pore channels on T cell cytolytic granules to 
            stimulate exocytosis and killing.
  JOURNAL   Curr Biol. 2012 Dec 18;22(24):2331-7. doi: 
            10.1016/j.cub.2012.10.035. Epub 2012 Nov 21.
  PUBMED    23177477
REFERENCE   2  (bases 1 to 6991)
  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"; doi: 
            "10.1016/j.cub.2012.10.035"; journalName: "Curr Biol"; date: 
            "2012-12-18- 18"; volume: "22"; issue: "24"; pages: "2331-7"
COMMENT     Sequence Label: HsTPC2-EYFP
FEATURES             Location/Qualifiers
     source          1..6991
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        75..378
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        379..582
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     533..553
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2945..2954
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2951..2953,2954..2956,2957..3670)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2996..3017)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3257..3276)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3604..3625
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3793..3914
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3830..3849)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3884..3903
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3921..4376)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4058..4079)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4270..4289
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4403..4507
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4509..4866
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4509..4700
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4532..4552)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4717..4852
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4901..5695
                     /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             4901..5695
                     /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(4955..4974)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5565..5584
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5883..5902)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5927..5974
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6303..6891
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6792..6811
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttaccgccat gcattagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
       61 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
      121 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      181 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      241 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      301 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      361 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      421 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      481 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      541 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag
      601 atccgctagc gctaccggac tcagatctcg agctcaagct tccatggcgg aaccccaggc
      661 ggagtcggag cccctgctgg gcggggcccg cggcggtggc ggcgactggc cggcggggct
      721 gaccacttac cgcagcatcc aagtcggccc tggtgccgcg gccaggtggg acctctgcat
      781 tgatcaggct gtggtcttca tcgaagatgc tattcagtac cgctccatca accaccgggt
      841 ggatgccagc tcgatgtggc tttaccgacg gtattactcg aacgtatgcc aacggacttt
      901 gagcttcacc atcttcttga tcctgttttt ggcttttatc gagaccccat cctcactcac
      961 cagcacggcg gacgtgcgct accgcgctgc cccctgggag ccgccctgcg gcctgaccga
     1021 gagtgtcgag gtgctctgcc tgctggtctt tgcggccgac ctctctgtga agggttacct
     1081 gttcgggtgg gcccatttcc agaaaaacct ttggctgctg ggctacctcg tggtgctggt
     1141 ggtgtctctg gtggactgga ccgtgtccct gagtctcgtg tgtcatgagc ccctgcggat
     1201 ccgccggctt ctccgtccct tcttcctgct gcagaactcc tctatgatga agaagacctt
     1261 gaaatgcatc cgctggtcgc tgccggaaat ggccagcgtc gggctgctgc tggccatcca
     1321 cctgtgcctc ttcaccatgt tcggaatgct gctgttcgct ggtgggaagc aggatgatgg
     1381 gcaggacagg gagaggctga cctacttcca gaacctgcct gagtctctga cttccctcct
     1441 ggtgctgctg accacggcca acaaccccga tgtgatgatt cctgcgtatt ccaagaaccg
     1501 ggcctatgcc atcttcttca tagtcttcac tgtgatagga agcctgtttc tgatgaacct
     1561 gctgacagcc atcatctaca gtcagttccg gggctacctg atgaaatctc tccagacctc
     1621 gctgtttcgg aggcggctgg gaacccgggc tgcctttgaa gtcctatcct ccatggtggg
     1681 ggagggagga gccttccctc aggcagttgg ggtgaagccc cagaacttgc tgcaggtgct
     1741 tcagaaggtc cagctggaca gctcccacaa acaggccatg atggagaagg tgcgttccta
     1801 cggcagtgtt ctgctgtcag ctgaggagtt tcagaagctc ttcaacgagc ttgacagaag
     1861 tgtggttaaa gagcacccgc cgaggcccga gtaccagtct ccgtttctgc agagcgccca
     1921 gttcctcttc ggccactact actttgacta cctggggaac ctcatcgccc tggcaaacct
     1981 ggtgtccatt tgcgtgttcc tggtgctgga tgcagatgtg ctgcctgctg agcgtgatga
     2041 cttcatcctg gggattctca actgcgtctt cattgtgtac tacctgttgg agatgctgct
     2101 caaggtcttt gccctgggcc tgcgagggta cctgtcctac cccagcaacg tgtttgacgg
     2161 gctcctcacc gttgtcctgc tggttttgga gatctcaact ctggctgtgt accgattgcc
     2221 acacccaggc tggaggccgg agatggtggg cctgctgtcg ctgtgggaca tgacccgcat
     2281 gctgaacatg ctcatcgtgt tccgcttcct gcgtatcatc cccagcatga agccgatggc
     2341 cgtggtggcc agtaccgtcc tgggcctggt gcagaacatg cgtgcttttg gcgggatcct
     2401 ggtggtggtc tactacgtat ttgccatcat tgggatcaac ttgtttagag gcgtcattgt
     2461 ggctcttcct ggaaacagca gcctggcccc tgccaatggc tcggcgccct gtgggagctt
     2521 cgagcagctg gagtactggg ccaacaactt cgatgacttt gcggctgccc tggtcactct
     2581 gtggaacttg atggtggtga acaactggca ggtgtttctg gatgcatatc ggcgctactc
     2641 aggcccgtgg tccaagatct attttgtatt gtggtggctg gtgtcgtctg tcatctgggt
     2701 caacctgttt ctggccctga ttctggagaa cttccttcac aagtgggacc cccgcagcca
     2761 cctgcagccc cttgctggga ccccagaggc cacctaccag atgactgtgg agctcctgtt
     2821 cagggatatt ctggaggagc ccgaggagga tgagctcaca gagaggctga gccagcaccc
     2881 gcacctgtgg ctgtgcagga gaattctgca gtcgacggta ccgcgggccc gggatccacc
     2941 ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     3001 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     3061 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     3121 ctggcccacc ctcgtgacca ccttcggcta cggcctgcag tgcttcgccc gctaccccga
     3181 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     3241 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     3301 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     3361 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     3421 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     3481 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     3541 cgacaaccac tacctgagct accagtccgc cctgagcaaa gaccccaacg agaagcgcga
     3601 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     3661 gtacaagtaa agcggccgcg actctagatc ataatcagcc ataccacatt tgtagaggtt
     3721 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     3781 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     3841 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     3901 atcaatgtat cttaaggcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa
     3961 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa
     4021 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact
     4081 attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc
     4141 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     4201 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     4261 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     4321 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg
     4381 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     4441 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     4501 gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag
     4561 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     4621 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     4681 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     4741 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     4801 gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     4861 tgcaaagatc gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     4921 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     4981 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     5041 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct
     5101 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     5161 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     5221 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     5281 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     5341 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     5401 agccgaactg ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac
     5461 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     5521 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     5581 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     5641 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     5701 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     5761 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     5821 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccctag ggggaggcta
     5881 actgaaacac ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga
     5941 cagaataaaa cgcacggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc
     6001 tggcactctg tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc
     6061 ttttccccac cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg
     6121 gcggcaggcc ctgccatagc ctcaggttac tcatatatac tttagattga tttaaaactt
     6181 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     6241 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     6301 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     6361 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     6421 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     6481 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     6541 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     6601 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     6661 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     6721 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     6781 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     6841 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     6901 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     6961 gcgttatccc ctgattctgt ggataaccgt a
//
