LOCUS       sequence-441729-        6848 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Biosensor for extracellular L-lactate.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6848)
  AUTHORS   Kamijo Y, Machler P, Ness N, Vu CQ, Kusakizako T, Mannuthodikayil J,
            Ku Z, Boisvert M, Grebenik E, Miyazaki I, Hashizume R, Sato H, Liu 
            R, Hori Y, Tomita T, Katayama T, Furube A, Caraveo G, Paquet ME, 
            Drobizhev M, Nureki O, Arai S, Brancaccio M, Campbell RE, Kleinfeld 
            D, Nasu Y
  TITLE     A red fluorescent genetically encoded biosensor for in vivo imaging 
            of extracellular L-lactate dynamics.
  JOURNAL   Nat Commun. 2025 Oct 29;16(1):9531. doi: 10.1038/s41467-025-64484-x.
  PUBMED    41162396
REFERENCE   2  (bases 1 to 6848)
  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: "Nat 
            Commun."; date: "2025-10-29"; volume: "16(1):9531. doi"; pages: " 
            10.1038/s41467-025-64484-x"
COMMENT     Sequence Label: pAAV-gfaABC1D-IgK-R-eLACCO2-COBRA
FEATURES             Location/Qualifiers
     source          1..6848
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     primer_bind     895..914
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        895..913
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      938..947
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     2293..2312
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    2975..3563
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3028..3048)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3446..3457)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3595..4071
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3794..3813)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4111..4251
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4111..4240
                     /label=AAV2 ITR
     rep_origin      4326..4781
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4413..4432)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4623..4644
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4798..4817)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4917..4939
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4977..4995)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5063..5167
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5168..5236,5237..6028)
                     /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"
     rep_origin      6199..6787
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6688..6707
                     /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 gcgtggatct aacatatcct ggtgtggagt aggggacgct
      181 gctctgacag aggctcgggg gcctgagctg gctctgtgag ctggggagga ggcagacagc
      241 caggccttgt ctgcaagcag acctggcagc attgggctgg ccgcccccca gggcctcctc
      301 ttcatgccca gtgaatgact caccttggca cagacacaat gttcggggtg ggcacagtgc
      361 ctgcttcccg ccgcacccca gcccccctca aatgccttcc gagaagccca ttgagcaggg
      421 ggcttgcatt gcaccccagc ctgacagcct ggcatcttgg gataaaagca gcacagcccc
      481 ctaggggctg cccttgctgt gtggcgccac cggcggtgga gaacaaggct ctattcagcc
      541 tgtgcccagg aaaggggatc aggggatgcc caggcatgga cagtgggtgg caggggggga
      601 gaggagggct gtctgcttcc cagaagtcca aggacacaaa tgggtgaggg gagagctctc
      661 cccatagctg ggctgcggcc caaccccacc ccctcaggct atgccagggg gtgttgccag
      721 gggcacccgg gcatcgccag tctagcccac tccttcataa agccctcgca tcccaggagc
      781 gagcagagcc agagcaggtt ggagaggaga cgcatcacct ccgctgctcg cggggatcca
      841 ccggtgagct ctctggctaa ctagagaacc cactgcttac tggcttatcg aaattaatac
      901 gactcactat agggagaccc aagctggcta gcgtgccgcc accatggcct cgagaatgga
      961 gaccgacacc ctgctgctgt gggtgctgct gctgtgggtg cccggctcca ccggcgacag
     1021 atctatgttc tctcccatgg cagtggccca ggctcgacgc tataagtgga ggattcaaac
     1081 cgcgtgggat gctggaacag taggctacag tctgttccaa aagtttacgg agcgggtaaa
     1141 ggaactgacg gatggacagc ttgaggtcca gcctcttcca gcgggtgcgg tcgtaggcac
     1201 gtttgatatg ttcgacgcgg tcaagactgg agttttggac ggaatgaatc ctttcacgct
     1261 gtattgggca gagaggatgc cggttacagc tttcctctcc agctacgccc tcgggttgga
     1321 ccgacctgac cagtgggaaa catggtttta ctccctgggt ggcctcgaca acgcccgacg
     1381 agcctttgcc gagcaaggcc tcttctacgt gggaccagtt cagcacgacc tcaatacaat
     1441 tcattcaaag aagccgatcc ggagatttga ggacttcaaa ggtgtcaagc tccgagtgcc
     1501 aggcggcatg attgcggaag tctttgcggc cgcaggagct tccaccgtcc tcttgcctgg
     1561 gggggaggtc taccccgctc tggaaagagg ggcagtgact gcggtttccg agcggatgta
     1621 ccccgaggac ggcgccctga agagcgagat caagaagggg ctgaggctga aggacggcgg
     1681 ccactacgcc gccgtggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg
     1741 cgcctacgtc gtcgacatcc agttggacat cgtgtcccac aacgaggact acactatcgt
     1801 ggaacagtgc gaacgcgccg agggccgcca ctccaccggc ggcgtggtcg agctggacaa
     1861 gggaggtaca ggcgggagtc tggtgagcaa gggcgaggag gatgacatgg ccatcgtcaa
     1921 ggagttcatg cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat
     1981 cgagggcgag ggcgagggcc gcccctacga ggcctttcag accgctaagc tgaaggtgac
     2041 caagggtggc cccctgccct tcgcctggga catcctatcc cctcagttca tgtacggctc
     2101 caaggcctac attaagcacc cagccgacat ccccgactac ttcaagctgt ccttccccga
     2161 gggcttcagg tgggagcgcg tgatgtactt cgaggacggc ggcattattc acgttgacca
     2221 ggactcctcc ctgcaggacg gcgtattcat ctacaaggtg aagctgcgcg gcaccaactt
     2281 cccccccgac ggccccgtaa tgcagaagaa gaccatgggc tgggagcggg ggcgtagtgc
     2341 ggccgatttt gttgggcctg cagtaaacta taacttgggg ttccatcagg aggcaaagta
     2401 catcataatg gggcctccag agacgcctgc cattcatcaa cctgtagacc ttatggactt
     2461 caccataaac cttaatcggt ggcgcagcct tccaaagcct ctgcaagaaa gattcatcgc
     2521 ggcggttcac gagtactctt ggattcacta cgcaggcatt cagaaggcta atttggaggc
     2581 gtggccgaaa tatcggcagg caggtgttga ggtcatccgg ctgggcaacg aggatgttag
     2641 aaaattcagg cgactcgcta tccccatttg gtttaagtgg gctaagatgg ataaatactc
     2701 acgagaggct ttcgcgtccc aactggagta tatgagaggt ataggtcacg ttacggacga
     2761 ggaattgaag ggcttgagcc tgggctccac ctccggctcc ggcaagcccg gctccggcga
     2821 gggctccacc aagggcgaat tctcagataa cgactacggt aaagatgggt tggctggggc
     2881 ttccggaggc agacgaaatg gcgctataac tgtgctcagt ttcataacgt tctacgtggc
     2941 agcatttatg gtattgttgt agaagcttat cgataatcaa cctctggatt acaaaatttg
     3001 tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc
     3061 tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta
     3121 taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt
     3181 ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca
     3241 gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc
     3301 ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt
     3361 gtcggggaaa tcatcgtcct ttccttggct gctcgcctat gttgccacct ggattctgcg
     3421 cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg
     3481 cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat
     3541 ctccctttgg gccgcctccc cgcatcgata ccgagcgctg ctcgagagat ctacgggtgg
     3601 catccctgtg acccctcccc agtgcctctc ctggccctgg aagttgccac tccagtgccc
     3661 accagccttg tcctaataaa attaagttgc atcattttgt ctgactaggt gtccttctat
     3721 aatattatgg ggtggagggg ggtggtatgg agcaaggggc aagttgggaa gacaacctgt
     3781 agggcctgcg gggtctattg ggaaccaagc tggagtgcag tggcacaatc ttggctcact
     3841 gcaatctccg cctcctgggt tcaagcgatt ctcctgcctc agcctcccga gttgttggga
     3901 ttccaggcat gcatgaccag gctcagctaa tttttgtttt tttggtagag acggggtttc
     3961 accatattgg ccaggctggt ctccaactcc taatctcagg tgatctaccc accttggcct
     4021 cccaaattgc tgggattaca ggcgtgaacc actgctccct tccctgtcct tctgattttg
     4081 taggtaacca cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc
     4141 cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg
     4201 gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga
     4261 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc
     4321 atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     4381 gaccgctaca cttgccagcg ccttagcgcc cgctcctttc gctttcttcc cttcctttct
     4441 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     4501 atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag
     4561 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     4621 tagtggactc ttgttccaaa ctggaacaac actcaactct atctcgggct attcttttga
     4681 tttataaggg attttgccga tttcggtcta ttggttaaaa aatgagctga tttaacaaaa
     4741 atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac
     4801 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     4861 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     4921 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     4981 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     5041 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     5101 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     5161 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     5221 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     5281 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     5341 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     5401 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     5461 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     5521 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     5581 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     5641 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     5701 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     5761 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     5821 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     5881 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     5941 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     6001 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     6061 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     6121 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     6181 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     6241 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     6301 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     6361 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     6421 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     6481 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     6541 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     6601 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     6661 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     6721 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     6781 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     6841 tcacatgt
//
