LOCUS       sequence-361926-        3877 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for lactate (with lifetime and intensity 
            response); for bacterial or in vitro expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3877)
  AUTHORS   Koveal D, Rosen PC, Meyer DJ, Diaz-Garcia CM, Wang Y, Cai LH, Chou 
            PJ, Weitz DA, Yellen G
  TITLE     A high-throughput multiparameter screen for accelerated development 
            and optimization of soluble genetically encoded fluorescent 
            biosensors.
  JOURNAL   Nat Commun. 2022 May 25;13(1):2919. doi: 10.1038/s41467-022-30685-x.
  PUBMED    35614105
REFERENCE   2  (bases 1 to 3877)
  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: "2022-05-25"; volume: "13(1):2919. doi"; pages: " 
            10.1038/s41467-022-30685-x"
COMMENT     Sequence Label: pRSetB-KanR-LiLac
FEATURES             Location/Qualifiers
     source          1..3877
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     489..508
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     742..759
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     844..863
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        844..862
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             894..916
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             930..950
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     primer_bind     complement(993..1014)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1254..1273)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             2223..2471
                     /codon_start=1
                     /product="C-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VC155
                     /translation="DKQKNGIKANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNH
                     YLSTQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELY"
     terminator      2486..2533
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     2623..2644
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2807..2911
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2912..3706
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2912..3706
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2966..2985)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3576..3595
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      join(3877,1..588)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
       61 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
      121 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
      181 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
      241 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
      301 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
      361 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
      421 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
      481 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
      541 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
      601 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
      661 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
      721 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata
      781 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcaggatctc gatcccgcga
      841 aattaatacg actcactata gggagaccac aacggtttcc ctctagaaat aattttgttt
      901 aactttaaga aggagatata catatgaaac atcaccatca tcatcatcat gtgagcaagg
      961 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
     1021 gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
     1081 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc
     1141 tgtcctgggg cgtgcagtgc ttcgcccgct accccgacca catgaagcag cacgacttct
     1201 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
     1261 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
     1321 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
     1381 actcctccgg ccgcaccggc attgacccct ttactgaaag cgtgttgcaa agccaagcca
     1441 cagaattgct gcaaaaaaaa gctcagttag tcagttttaa aattcaaggc attatgaagc
     1501 gcatttttat gggcgctaac acccttgaaa aatttttaag cgatgaaaat agcgctatca
     1561 acgacaccct caaacgccgc atgctctctg agtttttgtt agcaaaccct catgtgttat
     1621 tggttagcgc gatttatacg aataataatg aacgtgtcat cactgccatg agcatggatt
     1681 caaaaatcgc ctaccctaat accacgctca atgaaaacat gaccaatcaa atccgttcgc
     1741 tcaaaagtat tacccattca gatccctatt ataaagaggt taatggcgat aaaatctatg
     1801 gcatggatat taccctcccc cttatgggta agaatcaaaa tgctattggc gcgctgaatt
     1861 tctttttaaa cattgacgct ttttataccg atgtggtagg caagaaaaag agcaacacct
     1921 ttttaatggg gaaagacggc cgtcttttaa tcaaccctaa tcgtgagatc caagataaga
     1981 ttttaagcgc tatcaatcca gatcgccgtg tcgctaaagc tgtggagtat tacaatcaaa
     2041 acgaagcggg cactttgagc taccattcat tgagcgggaa tacagaaacc tttttagcca
     2101 ttcagccctt tgattttttt gaagaaaaag ggaataacgg caatcattgg cgttgggcaa
     2161 ttgggaaata tgtcaataaa tctttagtct tttccagcca cagcaacgtc tatatcaccg
     2221 ccgacaagca gaagaacggc atcaaggcca acttcaagat ccgccacaac atcgaggacg
     2281 gcggcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc
     2341 tgctgcccga caaccactac ctgagcaccc agtccaagct gagcaaagac cccaacgaga
     2401 agcgcgatca catggtccta ctggagttcg tgaccgccgc cgggatcact ctcggcatgg
     2461 acgagctgta ccagtgattt ccggactagc ataacccctt ggggcctcta aacgggtctt
     2521 gaggggtttt ttgctgagct ccgaggggcc caagctgtag tgggccatcg ccctgataga
     2581 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     2641 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     2701 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     2761 aaatattaac gcttacaatt taggtggcac ttttcgggga aatgtgcgcg gaacccctat
     2821 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     2881 aatgcttcaa taatattgaa aaaggaagag tatgattgaa caagatggat tgcacgcagg
     2941 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     3001 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     3061 gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct
     3121 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     3181 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     3241 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     3301 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     3361 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     3421 gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     3481 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     3541 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     3601 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     3661 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgactgt cagaccaagt
     3721 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     3781 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     3841 agcgtcagac cccgtagaaa agatcaaagg atcttct
//
