LOCUS       sequence-408944-        6631 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Biosensor for intracellular L-lactate.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6631)
  AUTHORS   Nasu Y, Aggarwal A, Le GNT, Vo CT, Kambe Y, Wang X, Beinlich FRM, 
            Lee AB, Ram TR, Wang F, Gorzo KA, Kamijo Y, Boisvert M, Nishinami S,
            Kawamura G, Ozawa T, Toda H, Gordon GR, Ge S, Hirase H, Nedergaard 
            M, Paquet ME, Drobizhev M, Podgorski K, Campbell RE
  TITLE     Lactate biosensors for spectrally and spatially multiplexed 
            fluorescence imaging.
  JOURNAL   Nat Commun. 2023 Oct 27;14(1):6598. doi: 10.1038/s41467-023-42230-5.
  PUBMED    37891202
REFERENCE   2  (bases 1 to 6631)
  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: "2023-10-27"; volume: "14(1):6598. doi"; pages: " 
            10.1038/s41467-023-42230-5"
COMMENT     Sequence Label: pcDNA-R-iLACCO1.2
FEATURES             Location/Qualifiers
     source          1..6631
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        110..489
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        490..693
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     644..664
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     738..757
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        738..756
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      781..790
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1821..1840
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2097..2114)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2103..2327
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2373..2801
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2460..2479)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2670..2691
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2810..2830)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2815..3144
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2995..3130
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3057..3076
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3211..4005
                     /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(3265..3284)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3875..3894
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4179..4300
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4216..4235)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4270..4289
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4349..4365)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4349..4365)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4362..4384)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4373..4389
                     /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(4397..4403,4404..4421,4422..4427))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4442..4463
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4580..4597)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4751..5339)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4831..4850)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5510..6301,6302..6370))
                     /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     6133..6152
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6371..6475)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6550..6569)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
       61 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
      121 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      181 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      241 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      301 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      361 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      421 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      481 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      541 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      601 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      661 cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga acccactgct
      721 tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg ctagcgtgcc
      781 gccaccatgg cctcgagaat ggaacaaaac atagtccagc cgttgaagac cctcatggga
      841 gacgaccctg attattcatt tgatattctg gaggcgcggt acgcgataga agcgtcaact
      901 gcctggtacg ctgcactgcg agctacaccc ggggacaaag agaaaattca actgtgcttc
      961 gaggctacac ggtccgaaga tcccgatata gcttcccaag cagacgtaag gttccacctg
     1021 gcgattgctg aagcaagcca taacatagtt ctgcttcaga caatgcgagg cttttttgat
     1081 gttttgcagt cttcagtcaa gcacagtcgg cagcggatgt acttggattg ggtttccgag
     1141 cggatgttcc ccgaggacgg cgccctgaag agcgagatca agaaggggct gaggctgaag
     1201 gacggcggcc actacgccgc cgaggtcaag accacttaca aggccaagaa gcccgtgcag
     1261 ctgcccggcg cctacgtcgt cgacatcaag ttggacatcg tgtcccacaa cgaggactac
     1321 actatcgtgg aacagtgcga acgcgccgag ggccgccact ccaccggcgg cgtggacgag
     1381 ctgtacaagg gaggtacagg cgggagtctg gtgagcaagg gcgaggagga tgacatggcc
     1441 atcgtcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtaaa cggccacgag
     1501 ttcgagatcg agggcgaggg cgagggccgc ccctacgagg cctttcagac cgctaagctg
     1561 aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctatcccc tcagttcatg
     1621 tacggctcca aggcctacat taagcaccca gccgacatcc ccgactactt caagctgtcc
     1681 ttccccgagg gcttcaggtg ggagcgcgtg atgtacttcg aggacggcgg cattattcac
     1741 gttaaccagg actcctccct gcaggacggc gtattcatct acaaggtgaa gctgcgcggc
     1801 accaacttcc cccccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcgagg
     1861 ccgccggtct acggtcaact cacagaacag catcaggcag ttatagacgc catctttgcc
     1921 ggcgatgcgg acggtgctag aaaagctatg atggctcatc tgagcttcat ccatacgact
     1981 atgaaacgct tcgatggaga tcaggcccga cacgcacgca tttccagact gcctggagag
     2041 cataatgagc actctcggga aaagaaagca tgaaagcttg gtaaaccgct gatcagcctc
     2101 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     2161 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     2221 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     2281 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     2341 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     2401 ggcgggtgtg gtggttacgc gcagtgtgac cgctacactt gccagcgccc tagcgcccgc
     2461 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     2521 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     2581 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     2641 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     2701 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     2761 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt
     2821 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     2881 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     2941 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     3001 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     3061 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt
     3121 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct
     3181 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
     3241 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
     3301 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
     3361 accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg
     3421 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
     3481 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
     3541 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
     3601 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
     3661 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
     3721 ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
     3781 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     3841 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     3901 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     3961 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     4021 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     4081 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     4141 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     4201 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     4261 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     4321 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     4381 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     4441 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     4501 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     4561 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     4621 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     4681 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     4741 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     4801 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     4861 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     4921 cttcgggaag cgtggcgctt tctcatagct cacgctatag gtatctcagt tcggtgtagg
     4981 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     5041 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     5101 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5161 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     5221 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5281 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     5341 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     5401 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     5461 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     5521 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     5581 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     5641 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     5701 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     5761 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     5821 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     5881 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     5941 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     6001 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     6061 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     6121 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     6181 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     6241 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     6301 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     6361 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     6421 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     6481 ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tcacccgatc ccctatggtg
     6541 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg
     6601 tgtgttggag gtcgctgagt agtgcgcgag c
//
