LOCUS       sequence-238528-        6821 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses LAR-GECO1.2 in the mitochondria in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6821)
  AUTHORS   Wu J, Prole DL, Shen Y, Lin Z, Gnanasekaran A, Liu Y, Chen L, Zhou 
            H, Chen SR, Usachev YM, Taylor CW, Campbell RE
  TITLE     Red fluorescent genetically encoded Ca2+ indicators for use in 
            mitochondria and endoplasmic reticulum.
  JOURNAL   Biochem J. 2014 Nov 15;464(1):13-22. doi: 10.1042/BJ20140931.
  PUBMED    25164254
REFERENCE   2  (bases 1 to 6821)
  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"; journalName: 
            "Biochem J"; date: "2014-11-15- 15"; volume: "464"; issue: "1"; 
            pages: "13-22"
COMMENT     Sequence Label: CMV-mito-LAR-GECO1.2
FEATURES             Location/Qualifiers
     source          1..6821
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        74..453
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        454..657
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     608..628
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     702..721
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        702..720
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             767..841
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             863..937
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     primer_bind     1738..1757
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2251..2268)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2257..2481
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2527..2955
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2614..2633)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2824..2845
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2964..2984)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2969..3298
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3149..3284
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3211..3230
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3365..4159
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3419..3438)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4029..4048
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4333..4454
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4370..4389)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4424..4443
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4503..4519)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4503..4519)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4516..4538)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4527..4543
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4551..4557,4558..4575,4576..4581))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4596..4617
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4734..4751)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4905..5493)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4985..5004)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5664..6455,6456..6524))
                     /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     6287..6306
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6525..6629)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6704..6723)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc
       61 agatatacgc gttgacattg attattgact agttattaat agtaatcaat tacggggtca
      121 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      181 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      241 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      301 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      361 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      421 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      481 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      541 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      601 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctctc
      661 tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac tcactatagg
      721 gagacccaag ctggctagcg tttaaacggg ccctctagac tcgagcatgt ctgttctgac
      781 tcctctgctg ctccggggtc tcacaggttc cgcaagaaga ctccccgtgc ctagggccaa
      841 aattcattca ctgggggacc ccatgagcgt gctcacccca ctcctgctgc gggggctgac
      901 cggcagcgct aggcggctgc cagtccccag ggccaagatc cacagtctcg gcgatcccaa
      961 ggatccaacc atggtcgact cttcacgtcg taagtggatt aagcgtggtc acgcagtcag
     1021 agctataggt cggctgagct cacccgtggt ttccgagcgg atgtaccccg aggacggcgc
     1081 tctgaagagc gagatcaaga aggggctgag gctgaaggac ggcggccact acgccgccga
     1141 ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct acatagtcga
     1201 catcaagttg gacatcgtgt cccacaacga ggactacacc atcgtggaac agtgcgaacg
     1261 cgccgtgggc cgccactcca ccggcggcat ggacgagctg tacaagggag gtacaggcgg
     1321 gagtctggtg agcaagggcg aggaggataa cagggccatc gtcaaggagt tcatgcgctt
     1381 caagttgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga
     1441 gggccgcccc tacgaggcct ttcagaccgc taagctgaag gtgaccaagg gtggccccct
     1501 gcccttcgca tgggacatcc tgtcccctca gttgatgtac ggctccaagg cctacattaa
     1561 gcacccagcc gacatccccg actacttcaa gctgtccttc cccgagggct tcaggtggga
     1621 gcgcgtgatg aacttcgagg acggcggcat tattcacgtt agccaggaca cctccctgca
     1681 ggacggcgta ttcatctaca aggtgaagct gcgcggcacc aacttccccc ccgacggccc
     1741 cgtaatgcag aagaagacca tgggctggga ggctacgcgt gaccaactga ctgaagagca
     1801 gatcgcagaa tttaaagagg ctttctccct atttgacgag gacggggatg ggacgatgac
     1861 aaccaaggag ctggggacgg tgatgcggtc tctggggcag aaccccacag aagcagagct
     1921 gcaggacatg atcaatgaag tagatgccga cggtgacggc acattcgact tccctgagtt
     1981 cctgacgatg atggcaagaa aaatgaatga cacagacagt gaagaggaaa ttagagaagc
     2041 gttccgcgtg tttgataagg acggcaatgg ctacatcggc gcagcagagc ttcgccacgt
     2101 gatgacagac cttggagaga agataacaga tgaggaggtt gatgaaatga tcagggtagc
     2161 agacatcgat ggggatggtc aggtaaacta cgaagagttt gtacaaatga tgacagcgaa
     2221 gtaaaagctt aagtttaaac cgctgatcag cctcgactgt gccttctagt tgccagccat
     2281 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     2341 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     2401 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg
     2461 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt
     2521 atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     2581 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     2641 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     2701 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     2761 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     2821 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     2881 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     2941 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     3001 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     3061 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     3121 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     3181 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     3241 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     3301 gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat gaggatcgtt
     3361 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     3421 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     3481 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     3541 actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     3601 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     3661 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     3721 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     3781 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     3841 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc
     3901 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     3961 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     4021 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     4081 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     4141 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     4201 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     4261 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     4321 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     4381 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     4441 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca
     4501 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     4561 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     4621 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     4681 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     4741 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     4801 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4861 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4921 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4981 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     5041 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     5101 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     5161 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     5221 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     5281 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     5341 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     5401 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     5461 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     5521 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     5581 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     5641 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     5701 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     5761 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5821 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5881 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5941 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     6001 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     6061 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     6121 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     6181 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     6241 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     6301 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     6361 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     6421 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     6481 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     6541 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     6601 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     6661 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
     6721 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
     6781 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga c
//
