LOCUS       sequence-420448-        6494 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  green fluorescent FLIM indicator for endogenous ATP imaging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6494)
  AUTHORS   Arai S, Itoh H, Vu CQ, Nakayama M, Oshima M, Morita A, Okamoto K, 
            Okuda S, Teranishi A, Osawa M, Tamura Y, Nonoyama S, Takuma M, Fujie
            T, Sarker SR, Sudhaharan T, Kiya T, Lane EB, Kitaguchi T
  TITLE     qMaLioffG: A single green fluorescent protein FLIM indicator 
            enabling quantitative imaging of endogenous ATP
  JOURNAL   bioRxiv 2023.08.29.555275
REFERENCE   2  (bases 1 to 6494)
  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: "bioRxiv 
            2023.08.29.555275"
COMMENT     Sequence Label: qMaLioffG
FEATURES             Location/Qualifiers
     source          1..6494
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(972..993)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1233..1252)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             1800..2051
                     /codon_start=1
                     /product="C-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VC155
                     /translation="DKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNH
                     YLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK"
     primer_bind     1988..2009
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2085..2102)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2091..2315
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2361..2789
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2448..2467)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2658..2679
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2798..2818)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2803..3132
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2983..3118
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3045..3064
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3199..3993
                     /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(3253..3272)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3863..3882
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4167..4288
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4204..4223)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4258..4277
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4337..4353)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4337..4353)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4350..4372)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4361..4377
                     /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(4385..4391,4392..4409,4410..4415))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4430..4451
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4568..4585)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4739..5327)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4819..4838)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5498..6289,6290..6358))
                     /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     6121..6140
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6359..6463)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga ctcgagatgg tgagcaaagg cgaggagctg ttcaccgggg
      961 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg
     1021 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     1081 gcaagctgcc cgtgccctgg cccaccctcg tgaccacctt cggctacggc ctgatgtgct
     1141 tcgcccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     1201 gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg
     1261 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca
     1321 aggaggacgg caacatcctg gggcacaagc tggagcttcg ccctatgaag accgtgaagg
     1381 tgaacatcac cacccccgac ggccccgtgt acgacgccga catcgagatg gtgagcgtgc
     1441 gggccgagag cggcgacctg ggcatcctgc ccggccacat ccccaccaag gcccccctga
     1501 agatcggcgc cgtgcggctg aagaaggacg gccagaccga gatggtggcc gtgagcggcg
     1561 gcaacgtgga ggtgcggccc gaccaggtga ccatcaccgc ccaggccgcc gagaccgccg
     1621 agggcatcga caaggagcgg gccgaggccg cccggcagcg ggcccaggag cggctgaaca
     1681 gccagagcga cgacaccgac atccggcggg ccgagctggc cctgcagcgg gccctgaacc
     1741 ggctggacgt ggccggcaag gcactggaat tcaacagcca caacgtctat atcatggccg
     1801 acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca
     1861 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     1921 tgcccgacaa ccactacctg agctaccagt ccgccctgag caaagacccc aacgagaagc
     1981 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     2041 agctgtacaa gtaattcgaa gcttaagttt aaaccgctga tcagcctcga ctgtgccttc
     2101 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     2161 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
     2221 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
     2281 tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg
     2341 gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     2401 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     2461 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     2521 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     2581 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     2641 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     2701 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     2761 gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt
     2821 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     2881 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     2941 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     3001 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     3061 gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     3121 ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca
     3181 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     3241 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     3301 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     3361 ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc
     3421 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     3481 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     3541 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     3601 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     3661 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     3721 aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     3781 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     3841 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     3901 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     3961 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     4021 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     4081 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     4141 tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat
     4201 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     4261 gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga
     4321 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     4381 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     4441 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     4501 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     4561 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     4621 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     4681 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     4741 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     4801 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     4861 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     4921 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     4981 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     5041 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     5101 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     5161 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     5221 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     5281 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     5341 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     5401 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     5461 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     5521 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     5581 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     5641 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     5701 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     5761 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     5821 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     5881 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     5941 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     6001 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     6061 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     6121 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     6181 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     6241 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     6301 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     6361 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     6421 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     6481 tgccacctga cgtc
//
