LOCUS       sequence-449821-        5825 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 5825)
  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 5825)
  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: pBAD-R-eLACCO2
FEATURES             Location/Qualifiers
     source          1..5825
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     208..227
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             331..348
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             352..384
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(388..396,397..411)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     primer_bind     1695..1714
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2226..2243)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2396..2482
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2574..2601
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2620..2711
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2712..2780,2781..3572)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2930..2949)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3743..4331
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4232..4251
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4485..4502
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4517..4657
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4583..4602)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4743..4765
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4921..5799)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
       61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
      121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
      181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
      241 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc
      301 taacaggagg aattaaccat ggggggttct catcatcatc atcatcatgg tatggctagc
      361 atgactggtg gacagcaaat gggtcgggat ctgtacgacg atgacgataa ggatccgagc
      421 tcgagaatgt tctctcccat ggcagtggcc caggctcgac gctataagtg gaggattcaa
      481 accgcgtggg atgctggaac agtaggctac agtctgttcc aaaagtttac ggagcgggta
      541 aaggaactga cggatggaca gcttgaggtc cagcctcttc cagcgggtgc ggtcgtaggc
      601 acgtttgata tgttcgacgc ggtcaagact ggagttttgg acggaatgaa tcctttcacg
      661 ctgtattggg cagagaggat gccggttaca gctttcctct ccagctacgc cctcgggttg
      721 gaccgacctg accagtggga aacatggttt tactccctgg gtggcctcga caacgcccga
      781 cgagcctttg ccgagcaagg cctcttctac gtgggaccag ttcagcacga cctcaataca
      841 attcattcaa agaagccgat ccggagattt gaggacttca aaggtgtcaa gctccgagtg
      901 ccaggcggca tgattgcgga agtctttgcg gccgcaggag cttccaccgt cctcttgcct
      961 gggggggagg tctaccccgc tctggaaaga ggggcagtga ctgcggtttc cgagcggatg
     1021 taccccgagg acggcgccct gaagagcgag atcaagaagg ggctgaggct gaaggacggc
     1081 ggccactacg ccgccgtggt caagaccacc tacaaggcca agaagcccgt gcagctgccc
     1141 ggcgcctacg tcgtcgacat ccagttggac atcgtgtccc acaacgagga ctacactatc
     1201 gtggaacagt gcgaacgcgc cgagggccgc cactccaccg gcggcgtggt cgagctggac
     1261 aagggaggta caggcgggag tctggtgagc aagggcgagg aggatgacat ggccatcgtc
     1321 aaggagttca tgcgcttcaa ggtgcacatg gagggctccg tgaacggcca cgagttcgag
     1381 atcgagggcg agggcgaggg ccgcccctac gaggcctttc agaccgctaa gctgaaggtg
     1441 accaagggtg gccccctgcc cttcgcctgg gacatcctat cccctcagtt catgtacggc
     1501 tccaaggcct acattaagca cccagccgac atccccgact acttcaagct gtccttcccc
     1561 gagggcttca ggtgggagcg cgtgatgtac ttcgaggacg gcggcattat tcacgttgac
     1621 caggactcct ccctgcagga cggcgtattc atctacaagg tgaagctgcg cggcaccaac
     1681 ttcccccccg acggccccgt aatgcagaag aagaccatgg gctgggagcg ggggcgtagt
     1741 gcggccgatt ttgttgggcc tgcagtaaac tataacttgg ggttccatca ggaggcaaag
     1801 tacatcataa tggggcctcc agagacgcct gccattcatc aacctgtaga ccttatggac
     1861 ttcaccataa accttaatcg gtggcgcagc cttccaaagc ctctgcaaga aagattcatc
     1921 gcggcggttc acgagtactc ttggattcac tacgcaggca ttcagaaggc taatttggag
     1981 gcgtggccga aatatcggca ggcaggtgtt gaggtcatcc ggctgggcaa cgaggatgtt
     2041 agaaaattca ggcgactcgc tatccccatt tggtttaagt gggctaagat ggataaatac
     2101 tcacgagagg ctttcgcgtc ccaactggag tatatgagag gtataggtca cgttacggac
     2161 gaggaattga agggcttgag cctgtgaaag cttggctgtt ttggcggatg agagaagatt
     2221 ttcagcctga tacagattaa atcagaacgc agaagcggtc tgataaaaca gaatttgcct
     2281 ggcggcagta gcgcggtggt cccacctgac cccatgccga actcagaagt gaaacgccgt
     2341 agcgccgatg gtagtgtggg gtctccccat gcgagagtag ggaactgcca ggcatcaaat
     2401 aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa
     2461 cgctctcctg agtaggacaa atccgccggg agcggatttg aacgttgcga agcaacggcc
     2521 cggagggtgg cgggcaggac gcccgccata aactgccagg catcaaatta agcagaaggc
     2581 catcctgacg gatggccttt ttgcgtttct acaaactctt ttgtttattt ttctaaatac
     2641 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     2701 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     2761 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     2821 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     2881 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     2941 cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     3001 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     3061 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     3121 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     3181 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     3241 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     3301 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     3361 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     3421 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     3481 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     3541 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     3601 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     3661 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     3721 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     3781 aaacaaaaca accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     3841 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     3901 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     3961 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     4021 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     4081 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     4141 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     4201 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     4261 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     4321 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     4381 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     4441 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     4501 aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     4561 accgcatatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtata
     4621 cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc caacacccgc
     4681 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     4741 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgaggcagca
     4801 gatcaattcg cgcgcgaagg cgaagcggca tgcataatgt gcctgtcaaa tggacgaagc
     4861 agggattctg caaaccctat gctactccgt caagccgtca attgtctgat tcgttaccaa
     4921 ttatgacaac ttgacggcta catcattcac tttttcttca caaccggcac ggaactcgct
     4981 cgggctggcc ccggtgcatt ttttaaatac ccgcgagaaa tagagttgat cgtcaaaacc
     5041 aacattgcga ccgacggtgg cgataggcat ccgggtggtg ctcaaaagca gcttcgcctg
     5101 gctgatacgt tggtcctcgc gccagcttaa gacgctaatc cctaactgct ggcggaaaag
     5161 atgtgacaga cgcgacggcg acaagcaaac atgctgtgcg acgctggcga tatcaaaatt
     5221 gctgtctgcc aggtgatcgc tgatgtactg acaagcctcg cgtacccgat tatccatcgg
     5281 tggatggagc gactcgttaa tcgcttccat gcgccgcagt aacaattgct caagcagatt
     5341 tatcgccagc agctccgaat agcgcccttc cccttgcccg gcgttaatga tttgcccaaa
     5401 caggtcgctg aaatgcggct ggtgcgcttc atccgggcga aagaaccccg tattggcaaa
     5461 tattgacggc cagttaagcc attcatgcca gtaggcgcgc ggacgaaagt aaacccactg
     5521 gtgataccat tcgcgagcct ccggatgacg accgtagtga tgaatctctc ctggcgggaa
     5581 cagcaaaata tcacccggtc ggcaaacaaa ttctcgtccc tgatttttca ccaccccctg
     5641 accgcgaatg gtgagattga gaatataacc tttcattccc agcggtcggt cgataaaaaa
     5701 atcgagataa ccgttggcct caatcggcgt taaacccgcc accagatggg cattaaacga
     5761 gtatcccggc agcaggggat cattttgcgc ttcagccata cttttcatac tcccgccatt
     5821 cagag
//
