LOCUS       sequence-455224-        7732 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing a cytosolic pH sensor (mTagBFP2-mNeonGreen) in 
            Saccharomyces cerevisiae.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7732)
  AUTHORS   Evans S, Tominaga M, Lu Z, Bandari NC, McDonnell L, Wang C, Speight 
            RE, Ishii J, Vickers CE, Peng B
  TITLE     Characterization of Fluorescent Reporters for Flow Cytometry-Based 
            Single-Cell Studies in Saccharomyces cerevisiae.
  JOURNAL   ACS Synth Biol. 2025 Sep 19;14(9):3804-3812. doi: 
            10.1021/acssynbio.5c00432. Epub 2025 Sep 10.
  PUBMED    40927980
REFERENCE   2  (bases 1 to 7732)
  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"; doi: 
            "10.1021/acssynbio.5c00432"; journalName: "ACS Synth Biol"; date: 
            "2025-09-19- 19"; volume: "14"; issue: "9"; pages: "3804-3812"
COMMENT     Sequence Label: pILFPZ1A9
FEATURES             Location/Qualifiers
     source          1..7732
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      729..999
                     /gene="S. cerevisae PGK1"
                     /label=PGK1 terminator
                     /note="transcription terminator for 3-phosphoglycerate 
                     kinase gene"
     primer_bind     complement(1264..1280)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1264..1280)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1277..1299)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1288..1304
                     /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(1312..1318,1319..1336,1337..1342))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1357..1378
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1495..1512)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1666..2254)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1746..1765)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2425..3216,3217..3285))
                     /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     3048..3067
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3286..3390)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3458..3476
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3514..3536)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3636..3655
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3849..3871
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3863..3880
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3864..3880
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        5184..5527
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     terminator      5920..6117
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    complement(6324..6357)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        6577..6990
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
ORIGIN
        1 atggtgtcta aaggtgaaga ggataacatg gcttctttgc cagctactca tgaattgcat
       61 attttcggtt ccatcaacgg tgttgatttc gatatggttg gtcaaggtac tggtaatcca
      121 aatgatggtt acgaagagtt gaacctgaaa tctacaaaag gtgacttgca attctcccca
      181 tggattttgg ttccacatat tggttacggt ttccaccaat atttgccata tccagatggt
      241 atgtctccat ttcaagctgc tatggttgat ggttctggtt accaagttca tagaaccatg
      301 caatttgaag atggtgcttc tttgaccgtc aactacagat atacttacga aggttcccat
      361 attaagggtg aagctcaagt aaaaggtaca ggttttccag ctgatggtcc agttatgact
      421 aattctttga ctgctgctga ttggtgcaga tccaagaaaa cttatccaaa cgataagacc
      481 atcatctcca cctttaagtg gtcttacact actggtaacg gtaaaagata cagatccact
      541 gctagaacta cttacacttt tgctaaacct atggctgcca actacttgaa gaatcaacct
      601 atgtacgttt tccgtaagac cgaattgaaa cactccaaaa ctgagctgaa cttcaaagaa
      661 tggcaaaagg ctttcactga tgttatgggt atggatgagt tgtacaagta ataaagatct
      721 gatcttaaat tgaattgaat tgaaatcgat agatcaattt ttttcttttc tctttcccca
      781 tcctttacgc taaaataata gtttatttta ttttttgaat attttttatt tatatacgta
      841 tatatagact attatttatc ttttaatgat tattaagatt tttattaaaa aaaaattcgc
      901 tcctctttta atgcctttat gcagtttttt tttcccattc gatatttcta tgttcgggtt
      961 cagcgtattt taagtttaat aactcgaaaa ttctgcgttc gttaaagctt tcgagaagga
     1021 tattatttac tagtaggact atttgcaaag ggaagggatg ctaaggtaga gggtgaacgt
     1081 tacagaaaag caggctggga agcatatttg agaagatgcg gccagcaaaa ctaaaaaact
     1141 gtattataag taaatgcatg tatactaaac tcacaaatta gagcttcaat ttaattatat
     1201 cagttattac ccgggaatct cggtcgtaat gatttaaatg catgcaagct tggcgtaatc
     1261 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     1321 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     1381 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     1441 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     1501 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     1561 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     1621 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     1681 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     1741 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     1801 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     1861 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     1921 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     1981 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     2041 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     2101 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     2161 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     2221 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     2281 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     2341 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     2401 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     2461 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     2521 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     2581 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     2641 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     2701 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     2761 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     2821 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     2881 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     2941 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     3001 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     3061 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     3121 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     3181 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     3241 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     3301 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     3361 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     3421 ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt
     3481 tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac
     3541 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
     3601 gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag
     3661 agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
     3721 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     3781 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
     3841 agggttttcc cagtcacgac gttgtaaaac gacggccagt gaattcattt aaattttttt
     3901 gattcggtaa tctccgaaca gaaggaagaa cgaaggaagg agcacagact tagattggta
     3961 tatatacgca tatgtagtgt tgaagaaaca tgaaattgcc cagtattctt aacccaactg
     4021 cacagaacaa aaacctgcag gaaacgaaga taaaacatgt ccacaaaatc atataccagt
     4081 agagctgaga ctcatgcaag tccggttgca tcgaaacttt tacgtttaat ggatgaaaag
     4141 aagaccaatt tgtgtgcttc tcttgacgtt cgttcgactg atgagctatt gaaacttgtt
     4201 gaaacgttgg gtccatacat ttgccttttg aaaacacacg ttgatatctt ggatgatttc
     4261 agttatgagg gtactgtcgt tccattgaaa gcattggcag agaaatacaa gttcttgata
     4321 tttgaggaca gaaaattcgc cgatatcggt aacacagtca aattacaata tacatcgggc
     4381 gtttaccgta tcgcagaatg gtctgatatc accaacgccc acggggttac tggtgctggt
     4441 attgttgctg gcttgaaaca aggtgcgcaa gaggtcacca aagaaccaag gggattattg
     4501 atgcttgctg aattgtcttc caagggttct ctagcacacg gtgaatatac taagggtacc
     4561 gttgatattg caaagagtga taaagatttc gttattgggt tcattgctca gaacgatatg
     4621 ggaggaagag aagaagggtt tgattggcta atcatgaccc caggtgtagg tttagacgac
     4681 aaaggcgatg cattgggtca gcagtacaga accgtcgacg aagttgtaag tggtggatca
     4741 gatatcatca ttgttggcag aggacttttc gccaagggta gagatcctaa ggttgaaggt
     4801 gaaagataca gaaatgctgg atgggaagcg taccaaaaga gaatcagcgc tccccattaa
     4861 ttatacagga aacttaatag aacaaatcac atatttaatc taatagccac ctgcattggc
     4921 acggtgcaac actacttcaa cttcatctta caaaaagatc acgggatctg ttgtattggg
     4981 atctctaggc tgaagcttcg tacgctgcag gtcgacaacc cttaatgttc ttcattctac
     5041 gcacggtcct tcaattcgct tggcggagat ctatgacaag agggtcgaac tcgcctaagt
     5101 cgtaattgag tcagttacgc tagggataac agggtaatat agctgtttag cttgcctcgt
     5161 ccccgccggg tcacccggcc agcgacatgg aggcccagaa taccctcctt gacagtcttg
     5221 acgtgcgcag ctcaggggca tgatgtgact gtcgcccgta catttagccc atacatcccc
     5281 atgtataatc atttgcatcc atacattttg atggccgcac ggcgcgaagc aaaaattacg
     5341 gctcctcgct gcagacctgc gagcagggaa acgctcccct cacagacgcg ttgaattgtc
     5401 cccacgccgc gcccctgtag agaaatataa aaggttagga tttgccactg aggttcttct
     5461 ttcatatact tccttttaaa atcttgctag gatacagttc tcacatcaca tccgaacata
     5521 aacaaccatg ggtatgaccg accaagcgac gcccaacctg ccatcacgag atttcgatcc
     5581 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5641 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccgggc tcgatcccct
     5701 cgcgagttgg ttcagctgct gcctgaggct ggacgacctc gcggagttct accggcagtg
     5761 caaatccgtc ggcatccagg aaaccagcag cggctatccg cgcatccatg cccccgaact
     5821 gcaggagtgg ggaggcacga tggccgcttt ggtcgacccg gacgggacgc tcctgcgcct
     5881 gatacagaac gaattgcttg caggcatctc atgatcagta ctgacaataa aaagattctt
     5941 gttttcaaga acttgtcatt tgtatagttt ttttatattg tagttgttct attttaatca
     6001 aatgttagcg tgatttatat tttttttcgc ctcgacatca tctgcccaga tgcgaagtta
     6061 agtgcgcaga aagtaatatc atgcgtcaat cgtatgtgaa tgctggtcgc tatactgctg
     6121 tcgattcgat actaacgccg ccatccagtg tcgaaaacga gctctcgaga acccttaaag
     6181 ttacgctagg gataacaggg taatataggt tcttcattct acgcacggtc cttcaattcg
     6241 cttggcggtc tagaatgaca agagggtcga actcgcctaa gtcgtaattg agtccagatc
     6301 cactagtggc ctatctagac ctaataactt cgtatagcat acattatacg aagttatatt
     6361 aagggttgct cgagaaagag ctccagaaag cgaccaccca actttggctg ataatagcgt
     6421 ataaacaatg catactttgt acgttcaaaa tacaatgcag tagatatatt tatgcatatt
     6481 acatataata catatcacat aggaagcaac aggcgcgttg gacttttaat tttcgaggac
     6541 cgcgaatcct tacatcacac ccaatccccc acaagtgatc ccccacacac catagcttca
     6601 aaatgtttct actccttttt tactcttcca gattttctcg gactccgcgc atcgccgtac
     6661 cacttcaaaa cacccaagca cagcatacta aatttcccct ctttcttcct ctagggtgtc
     6721 gttaattacc cgtactaaag gtttggaaaa gaaaaaagag accgcctcgt ttctttttct
     6781 tcgtcgaaaa aggcaataaa aatttttatc acgtttcttt ttcttgaaaa tttttttttt
     6841 tgattttttt ctctttcgat gacctcccat tgatatttaa gttaataaac ggtcttcaat
     6901 ttctcaagtt tcagtttcat ttttcttgtt ctattacaac tttttttact tcttgctcat
     6961 tagaaagaaa gcatagcaat ctaatctaag ttttaattac aaactcgaga aaggatccaa
     7021 aatggtgtct aagggtgaag agctgatcaa agaaaacatg cacatgaagt tgtacatgga
     7081 aggtactgtt gataaccatc atttcaagtg tacctctgaa ggtgaaggta aaccatacga
     7141 aggtactcaa actatgagaa tcaaggttgt tgaaggtggt ccattgccat ttgcttttga
     7201 tattttggct acctccttct tgtacggttc taagactttc atcaaccaca ctcaaggtat
     7261 cccagatttc ttcaagcaat cttttccaga aggtttcact tgggaaagag ttactactta
     7321 cgaagatggt ggtgttttga ctgctactca agatacttct ttacaagacg gttgcttgat
     7381 ctacaacgtt aagattagag gtgtcaactt cacttctaac ggtccagtta tgcaaaagaa
     7441 aactttaggt tgggaagcct tcaccgaaac attatatcca gctgatggtg gtttggaagg
     7501 tagaaatgat atggctttga aattggttgg tggctcccat ttgattgcta acgctaagac
     7561 tacctacagg tctaagaagc cagctaagaa tttgaaaatg ccaggtgttt actacgtgga
     7621 ctatagattg gaaagaatca aagaagccaa caacgaaacc tacgttgaac aacacgaagt
     7681 tgctgttgct agatattgtg atttgccatc aaagttgggt cacaagttga ac
//
