LOCUS       sequence-244047-        6473 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Highly resposive GFP-based pyruvate nanosensor to explore 
            mitochondrial metabolism..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6473)
  AUTHORS   Arce-Molina R, Cortes-Molina F, Sandoval PY, Galaz A, Alegria K, 
            Schirmeier S, Barros LF, San Martin A
  TITLE     A highly responsive pyruvate sensor reveals pathway-regulatory role 
            of the mitochondrial pyruvate carrier MPC.
  JOURNAL   eLife. 2020 Mar 6;9. pii: 53917. doi: 10.7554/eLife.53917.
  PUBMED    32142409
REFERENCE   2  (bases 1 to 6473)
  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: "eLife."; 
            date: "2020-03-6"; volume: "9. pii: 53917. doi"; pages: " 
            10.7554/eLife.53917"
COMMENT     Sequence Label: Mito-PyronicSF/pCMV-myc-mito
FEATURES             Location/Qualifiers
     source          1..6473
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        12..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..593
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     544..564
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             636..710
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             2265..2294
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(2330..2347)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2336..2560
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2606..3034
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2693..2712)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2903..2924
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3043..3063)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3048..3377
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3228..3363
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3290..3309
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3403..4197
                     /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="MVEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3457..3476)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4067..4086
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4371..4492
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4408..4427)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4462..4481
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4525..4542)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4696..5284)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4776..4795)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5455..6246,6247..6315))
                     /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     6078..6097
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6316..6420)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtaccgaatt cacattgatt attgagtagt tattaatagt aatcaattac ggggtcatta
       61 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
      121 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      181 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
      241 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      301 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      361 atctacggtt agtcatcgct attaccatag tgatgcggtt ttggcagtac atcaatgggc
      421 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      481 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      541 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctttctggc
      601 taactagaga acccactgct tactggcacg tgaccatgtc cgtcctgacg ccgctgctgc
      661 tgcggggctt gacaggctcg gcccggcggc tcccagtgcc gcgcgccaag atccattcgt
      721 tgctgcaggt cgacctcgag atggcatata gcaaaattcg gcagcccaaa ctgagcgatg
      781 tgattgagca gcagctggag tttctgattc tggaaggcac cctgagacct ggagagaaac
      841 tgccccctga gcgcgaactc gccaagcagt tcgacgtgag tcgaccatca ctgagggagg
      901 ctatccagag gctggaagca aagggactgc tcctgaggag acagggagga gggactttcg
      961 tgcagagctc cctgtggcag agcttcagcg accccctggt cgagctcctg tctgaccacc
     1021 cagaaagtca gtacgatctc ctggagacaa gacatgctct ggaaggcatc gccgcttact
     1081 atgcagccct gcggtccact gacgaggata aggaacgcat ccgagagctg caccatgcca
     1141 ttgaactcgc tcagcagtca ggagacctgg atgcagagag caacgccgtg ctgcagtacc
     1201 agattgcagt caccgaggct gcacacaatg tggtcctcct gcatctcctg aggtgcatgg
     1261 agccaatgct ggcccagaac gtgagacaga attttgagct cctgctcgaa aacgtctata
     1321 tcaaggccga caagcagaaa aacggcatta aggctaactt caagatcaga cacaacatcg
     1381 aggatggtgg cgtgcagctg gcctaccatt atcagcagaa cacaccaatc ggagacggac
     1441 cagtgctgct cccagataat cactacctga gcgtccagtc caagctgtct aaagacccta
     1501 acgagaagcg ggatcatatg gtgctgctcg aatttgtcac agccgctggg atcactctgg
     1561 gtatggacga gctctataaa ggagggaccg gtggcagtat ggtgtcaaag ggcgaggaac
     1621 tgttcacagg agtggtcccc attctggtgg agctcgacgg cgatgtcaat ggacacaaat
     1681 tttccgtgtc tggcgagggc gaaggagatg ctacctacgg gaagctgaca ctcaaattca
     1741 tctgcaccac aggcaagctg ccagtgccct ggcctactct ggtcactacc ctcacctacg
     1801 gggtgcagtg tttctccaga tatcccgacc acatgaagca gcatgatttc tttaaatctg
     1861 ctatgcctga ggggtacatc caggaacgga caattttctt taaggacgat ggtaactaca
     1921 aaacacgcgc agaggtgaag ttcgaaggcg acactctggt caatcgaatc gagctgaagg
     1981 gaattgactt taaagaagat gggaacatcc tgggtcacaa gctggagtac aatactaggt
     2041 attctcggcg cgaaatgctg ccactcgtgt ctagtcacag gaccagaatc tttgaggcaa
     2101 ttatggccgg aaagcccgag gaagctagag aagcaagtca ccggcatctg gccttcatcg
     2161 aggaaattct gctcgaccgg agccgcgagg aatcccgaag ggagcgcagc ctgagacgac
     2221 tcgaacagcg aaagaactga ctcgagatca aacgggcggc cgcagaacaa aaactcatct
     2281 cagaagagga tctgaatggg gccgcatagt ctagaagctc gctgatcagc ctcgactgtg
     2341 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     2401 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     2461 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     2521 gacaatagca ggcatgctgg ggatggcccg ggctctatgg cttctgaggc ggaaagaacc
     2581 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     2641 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     2701 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     2761 gggctccctt tagggttccg atttagtgct ttacggcacc tctcccccaa aaaacttgat
     2821 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     2881 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     2941 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     3001 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
     3061 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     3121 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     3181 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     3241 ctccgcccag ttccgcccat tctccgcccc taggctgact aatttttttt atttatgcag
     3301 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     3361 gcctaggctt ttgcaaaaag ctcccccccc gggaggtcca caatggttga acaagatgga
     3421 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     3481 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     3541 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tccaggacga ggcagcgcgg
     3601 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     3661 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     3721 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     3781 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     3841 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     3901 ccagccgaac tgttcgccag gctcaaggcg cgtatgcccg acggcgagga tctcgtcgtg
     3961 actcatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     4021 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     4081 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     4141 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     4201 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     4261 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     4321 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     4381 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     4441 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     4501 gtataccgtc gatctttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4561 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4621 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     4681 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     4741 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     4801 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     4861 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     4921 gtaggtcgtt cgctccaagc tgggctgtat gcacgaaccc cccgttcagc ccgaccgctg
     4981 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     5041 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     5101 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     5161 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     5221 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5281 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5341 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5401 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5461 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5521 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     5581 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     5641 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     5701 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     5761 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     5821 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     5881 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     5941 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     6001 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     6061 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     6121 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     6181 gatgtaaccc actcgggcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     6241 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6301 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6361 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6421 cacatttccc cgaaaagtgc cacctgacgt cagatcgacg gatcgggaga tcg
//
