LOCUS       sequence-355965-        7725 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the FRET-based mitochondrial calcium indicator 4mtD3cpv 
            under control of the CaMKIIa promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7725)
  AUTHORS   Hagenston AM, Yan J, Bas-Orth C, Tan Y, Sekler I, Bading H
  TITLE     Disrupted expression of mitochondrial NCLX sensitizes neuroglial 
            networks to excitotoxic stimuli and renders synaptic activity toxic.
  JOURNAL   J Biol Chem. 2021 Dec 20:101508. doi: 10.1016/j.jbc.2021.101508.
  PUBMED    34942149
REFERENCE   2  (bases 1 to 7725)
  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 Biol 
            Chem. 2021 Dec 20:101508. doi: 10.1016/j.jbc.2021.101508."
COMMENT     Sequence Label: pAAV-4mtD3cpv
FEATURES             Location/Qualifiers
     source          1..7725
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        147..1435
                     /label=CaMKII promoter
                     /note="promoter for the mouse calcium/calmodulin-dependent 
                     kinase II alpha subunit"
     CDS             1478..1552
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             1574..1648
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             1670..1744
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             1766..1840
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     regulatory      1862..1871
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1868..1870,1871..1873,1874..2386)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     AHFKIRHNIE"
     CDS             3092..3826
                     /codon_start=1
                     /product="circularly permuted form of the Venus yellow 
                     fluorescent protein (Nagai et al., 2004)"
                     /label=cp173Venus
                     /note="efficient FRET acceptor for CFP"
                     /translation="MDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRD
                     HMVLLEFVTAAGITLGMDELYKGGSGGMVSKGEELFTGVVPILVELDGDVNGHKFSVSG
                     EGEGDATYGKLTLKLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPE
                     GYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHN
                     VYITADKQKNGIKANFKIRHNIE"
     misc_feature    3854..4442
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3907..3927)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4325..4336)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4445..4461)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4446..4462)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    4472..4948
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4671..4690)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4988..5128
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4988..5117
                     /label=AAV2 ITR
     rep_origin      5203..5658
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5290..5309)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5500..5521
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5675..5694)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5794..5816
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5854..5872)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5940..6044
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6045..6113,6114..6905)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6263..6282)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7076..7664
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7565..7584
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtaacatt atggccttag gtcacttcat ctccatgggg
      181 ttcttcttct gattttctag aaaatgagat gggggtgcag agagcttcct cagtgacctg
      241 cccagggtca catcagaaat gtcagagcta gaacttgaac tcagattact aatcttaaat
      301 tccatgcctt gggggcatgc aagtacgata tacagaagga gtgaactcat tagggcagat
      361 gaccaatgag tttaggaaag aagagtccag ggcagggtac atctacacca cccgcccagc
      421 cctgggtgag tccagccacg ttcacctcat tatagttgcc tctctccagt cctaccttga
      481 cgggaagcac aagcagaaac tgggacagga gccccaggag accaaatctt catggtccct
      541 ctgggaggat gggtggggag agctgtggca gaggcctcag gaggggccct gctgctcagt
      601 ggtgacagat aggggtgaga aagcagacag agtcattccg tcagcattct gggtctgttt
      661 ggtacttctt ctcacgctaa ggtggcggtg tgatatgcac aatggctaaa aagcagggag
      721 agctggaaag aaacaaggac agagacagag gccaagtcaa ccagaccaat tcccagagga
      781 agcaaagaaa ccattacaga gactacaagg gggaagggaa ggagagatga attagcttcc
      841 cctgtaaacc ttagaaccca gctgttgcca gggcaacggg gcaatacctg tctcttcaga
      901 ggagatgaag ttgccagggt aactacatcc tgtctttctc aaggaccatc ccagaatgtg
      961 gcacccacta gccgttacca tagcaactgc ctctttgccc cacttaatcc catcccgtct
     1021 gttaaaaggg ccctatagtt ggaggtgggg gaggtaggaa gagcgatgat cacttgtgga
     1081 ctaagtttgt tcgcatcccc ttctccaacc ccctcagtac atcaccctgg gggaacaggg
     1141 tccacttgct cctgggccca cacagtcctg cagtattgtg tatataaggc cagggcaaag
     1201 aggagcaggt tttaaagtga aaggcaggca ggtgttgggg aggcagttac cggggcaacg
     1261 ggaacagggc gtttcggagg tggttgccat ggggacctgg atgctgacga aggctcgcga
     1321 ggctgtgagc agccacagtg ccctgctcag aagccccaag ctcgtcagtc aagccggttc
     1381 tccgtttgca ctcaggagca cgggcaggcg agtggcccct agttctgggg gcagcggggg
     1441 atccaccggt cgactcacta tagggagacc caagcttatg tccgtcctga cgccgctgct
     1501 gctgcggggc ttgacaggct cggcccggcg gctcccagtg ccgcgcgcca agatccattc
     1561 gttgggggat cccatgtccg tcctgacgcc gctgctgctg cggggcttga caggctcggc
     1621 ccggcggctc ccagtgccgc gcgccaagat ccattcgttg gggaagctta tgtccgtcct
     1681 gacgccgctg ctgctgcggg gcttgacagg ctcggcccgg cggctcccag tgccgcgcgc
     1741 caagatccat tcgttggggg atcccatgtc cgtcctgacg ccgctgctgc tgcggggctt
     1801 gacaggctcg gcccggcggc tcccagtgcc gcgcgccaag atccattcgt tggggaagct
     1861 tgccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     1921 gctggacggc gacgtaaacg gccacaggtt cagcgtgtcc ggcgagggcg agggcgatgc
     1981 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     2041 gcccaccctc gtgaccaccc tgacctgggg cgtgcagtgc ttcagccgct accccgacca
     2101 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgtac
     2161 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     2221 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     2281 ggggcacaag ctggagtaca actacatcag ccacaacgtc tatatcaccg ccgacaagca
     2341 gaagaacggc atcaaggccc acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     2401 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     2461 caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     2521 catggtcctg ctggagttcg tgaccgccgc ccgcatgcat gaccaactga cagaagagca
     2581 gattgcagag ttcaaagaag ccttctcatt attggacaag gatggggacg gcaccatcac
     2641 cacaaaggaa cttggcaccg ctctgaggtc gcttggacaa aacccaacgg aagcagaatt
     2701 gcaggatatg atcaatgaag tcgatgctga tggcaatgga acgatttact ttcctgaatt
     2761 tcttactatg atggctagaa aaatgaagga cacagacagc gaagaggaaa tccgagaagc
     2821 attccgtgtt tttgacaagg atgggaacgg ctacatcagc gctgctgaat tacgtcacgt
     2881 catgacaaac ctcggggaga agttaacaga tgaagaagtt gatgaaatga taagggaagc
     2941 agatatcgat ggtgatggcc aagtaaacta tgaagagttt gtacaaatga tgacagcaaa
     3001 gggggggaag aggcgctggc agaaaacagg ccatgctgtc cgagcattcg gaagactgaa
     3061 gaagatctcc agctccgggg cactggagct catggacggc ggcgtgcagc tcgccgacca
     3121 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     3181 gagctaccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     3241 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agggtggcag
     3301 cggtggcatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     3361 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     3421 cacctacggc aagctgaccc tgaagctgat ctgcaccacc ggcaagctgc ccgtgccctg
     3481 gcccaccctc gtgaccaccc tgggctacgg cctgcagtgc ttcgcccgct accccgacca
     3541 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     3601 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     3661 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     3721 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcaccg ccgacaagca
     3781 gaagaacggc atcaaggcca acttcaagat ccgccacaac atcgagtaag aattcgatat
     3841 caagcttatc gataatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     3901 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     3961 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct
     4021 ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga
     4081 cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc
     4141 tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac
     4201 aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaaat catcgtcctt
     4261 tccttggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt
     4321 cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc
     4381 tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc
     4441 gcatcgatac cgtcgacctc gagagatcta cgggtggcat ccctgtgacc cctccccagt
     4501 gcctctcctg gccctggaag ttgccactcc agtgcccacc agccttgtcc taataaaatt
     4561 aagttgcatc attttgtctg actaggtgtc cttctataat attatggggt ggaggggggt
     4621 ggtatggagc aaggggcaag ttgggaagac aacctgtagg gcctgcgggg tctattggga
     4681 accaagctgg agtgcagtgg cacaatcttg gctcactgca atctccgcct cctgggttca
     4741 agcgattctc ctgcctcagc ctcccgagtt gttgggattc caggcatgca tgaccaggct
     4801 cagctaattt ttgttttttt ggtagagacg gggtttcacc atattggcca ggctggtctc
     4861 caactcctaa tctcaggtga tctacccacc ttggcctccc aaattgctgg gattacaggc
     4921 gtgaaccact gctcccttcc ctgtccttct gattttgtag gtaaccacgt gcggaccgag
     4981 cggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     5041 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     5101 agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat
     5161 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg
     5221 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct
     5281 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     5341 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     5401 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     5461 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     5521 gaacaacact caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt
     5581 cggtctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     5641 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     5701 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     5761 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     5821 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg
     5881 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     5941 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     6001 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     6061 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     6121 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     6181 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     6241 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     6301 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     6361 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     6421 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     6481 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     6541 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     6601 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     6661 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     6721 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     6781 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     6841 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     6901 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     6961 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     7021 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     7081 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     7141 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     7201 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     7261 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     7321 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     7381 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     7441 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     7501 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     7561 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     7621 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     7681 cctttttacg gttcctggcc ttttgctggc cttttgctca catgt
//
