LOCUS       sequence-247766-        8051 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  super Ecliptic pHluorin with C terminal mRuby2 fusion tag in yeast 
            expression vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8051)
  AUTHORS   Dodd BJT, Kralj JM
  TITLE     Live Cell Imaging Reveals pH Oscillations in Saccharomyces 
            cerevisiae During Metabolic Transitions.
  JOURNAL   Sci Rep. 2017 Oct 24;7(1):13922. doi: 10.1038/s41598-017-14382-0.
  PUBMED    29066766
REFERENCE   2  (bases 1 to 8051)
  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: "Sci Rep."; 
            date: "2017-10-24"; volume: "7(1):13922. doi"; pages: " 
            10.1038/s41598-017-14382-0"
COMMENT     Sequence Label: pKL06
FEATURES             Location/Qualifiers
     source          1..8051
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(11..29)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    71..574
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        611..715
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(716..784,785..1576)
                     /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     complement(934..953)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1747..2335
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2236..2255
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2489..2506
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2623..2644
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2659..2664,2665..2682,2683..2689)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2697..2713
                     /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)."
     primer_bind     2702..2724
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2721..2737
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2721..2737
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2756..2776
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2758..2776
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2794..3192
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     terminator      4680..4927
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4696..4714)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4945..4964)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4946..4964)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4974..4991)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4974..4990)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4983..5005)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5131..5586
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5218..5237)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5428..5449
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5887..6294
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             6295..7389
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     primer_bind     complement(7883..7902)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8002..8024
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt
       61 tcttaggacg gatcgcttgc ctgtaactta cacgcgcctc gtatctttta atgatggaat
      121 aatttgggaa tttactctgt gtttatttat ttttatgttt tgtatttgga ttttagaaag
      181 taaataaaga aggtagaaga gttacggaat gaagaaaaaa aaataaacaa aggtttaaaa
      241 aatttcaaca aaaagcgtac tttacatata tatttattag acaagaaaag cagattaaat
      301 agatatacat tcgattaacg ataagtaaaa tgtaaaatca caggattttc gtgtgtggtc
      361 ttctacacag acaagatgaa acaattcggc attaatacct gagagcagga agagcaagat
      421 aaaaggtagt atttgttggc gatcccccta gagtctttta catcttcgga aaacaaaaac
      481 tattttttct ttaatttctt tttttacttt ctatttttaa tttatatatt tatattaaaa
      541 aatttaaatt ataattattt ttatagcacg tgatgaaaag gacccaggtg gcacttttcg
      601 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
      661 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
      721 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
      781 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
      841 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
      901 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
      961 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     1021 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     1081 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     1141 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     1201 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     1261 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     1321 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     1381 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     1441 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     1501 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     1561 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     1621 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     1681 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     1741 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     1801 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     1861 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     1921 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     1981 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     2041 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     2101 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     2161 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     2221 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     2281 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     2341 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     2401 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     2461 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     2521 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     2581 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac
     2641 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
     2701 gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca agcgcgcaat
     2761 taaccctcac taaagggaac aaaagctgga gctcatagct tcaaaatgtt tctactcctt
     2821 ttttactctt ccagattttc tcggactccg cgcatcgccg taccacttca aaacacccaa
     2881 gcacagcata ctaaatttcc cctctttctt cctctagggt gtcgttaatt acccgtacta
     2941 aaggtttgga aaagaaaaaa gagaccgcct cgtttctttt tcttcgtcga aaaaggcaat
     3001 aaaaattttt atcacgtttc tttttcttga aaattttttt tttgattttt ttctctttcg
     3061 atgacctccc attgatattt aagttaataa acggtcttca atttctcaag tttcagtttc
     3121 atttttcttg ttctattaca acttttttta cttcttgctc attagaaaga aagcatagca
     3181 atctaatcta agttttctag agtaaaacac caagaactta gtttcgatgt ctaagggcga
     3241 ggaactgttc actggtgtgg ttccaatcct ggttgagctg gacggcgatg taaacggtca
     3301 caagttcagc gttagcggcg aaggtgaggg cgacgctacc tacggtaaac tgactctgaa
     3361 gttcatctgc accaccggta aactgcctgt tccgtggcca actctggtca ccaccttcag
     3421 ctacggtgtt cagtgtctga cccgctaccc tgatcacatg aaacgccatg acttcttcaa
     3481 aagcgcgatg ccggaaggct atgttcaaga acgtaccatc tttttcaagg atgacggcaa
     3541 ttataaaact cgtgccgaag taaagttcga aggcgacacc ctggttaacc gtattgaact
     3601 gaaaggtatt gatttcaaag aagatggtaa cattctgggt cacaaactgg aatataacta
     3661 caacgaccac caggtttaca tcatggcgga taaacagaaa aacggcatca aagctaactt
     3721 caaaatccgt cataacatcg aagacggcgg tgtgcagctg gctgaccact accagcagaa
     3781 caccccgatc ggtgatggtc cggttctgct gccggataat cactacctgt tcaccacgtc
     3841 tactctgtcc aaagacccga acgaaaaacg tgatcacatg gtcctgctgg aattcgttac
     3901 tgcagcaggt atcacccacg gcatggatga actgtacaag atggtttcaa agggtgaaga
     3961 acttattaaa gaaaacatga gaatgaaggt agttatggaa ggaagcgtta atggccatca
     4021 attcaagtgt acgggagaag gagagggtag gccttacgaa gggacgcaaa ctatgaggat
     4081 taaggttatt gagggtggtc cattgccatt cgcattcgac atcttagcta catcttttat
     4141 gtatggaagt cgtacgttca taaagtatcc tgctgatatt ccagacttct ttaaacagag
     4201 cttccctgag ggctttacat gggaaagagt gaccagatat gaagatggtg gtgttgttac
     4261 tgttactcaa gacacttctt tagaagacgg tgaactagtt tataatgtaa aagttagggg
     4321 tgttaacttc ccgtctaacg ggccagtcat gcaaaagaag acaaaaggat gggaacctaa
     4381 cacggagatg atgtatcccg ctgacggtgg tttgagaggc tatacggata ttgcgttgaa
     4441 agtcgacgga ggcggtcatt tgcattgcaa ttttgttaca acttacagga gtaagaagac
     4501 agtaggtaat atcaagatgc ccggcgttca cgcagttgac catcgtttgg aacgtattga
     4561 ggaatcagat aatgagacat acgtggtaca gagggaggtt gccgtagcta agtacagtaa
     4621 cttggggggt gggatggacg aattatataa ataaagctta tcgataccgt cgactcgagt
     4681 catgtaatta gttatgtcac gcttacattc acgccctccc cccacatccg ctctaaccga
     4741 aaaggaagga gttagacaac ctgaagtcta ggtccctatt tattttttta tagttatgtt
     4801 agtattaaga acgttattta tatttcaaat ttttcttttt tttctgtaca gacgcgtgta
     4861 cgcatgtaac attatactga aaaccttgct tgagaaggtt ttgggacgct cgaaggcttt
     4921 aatttgcggc cggtacccaa ttcgccctat agtgagtcgt attacgcgcg ctcactggcc
     4981 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca
     5041 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc
     5101 caacagttgc gcagcctgaa tggcgaatgg acgcgccctg tagcggcgca ttaagcgcgg
     5161 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     5221 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     5281 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     5341 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     5401 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     5461 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     5521 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     5581 caatttcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatatc
     5641 gaccggtcga ggagaacttc tagtatatct acatacctaa tattattgcc ttattaaaaa
     5701 tggaatccca acaattacat caaaatccac attctcttca aaatcaattg tcctgtactt
     5761 ccttgttcat gtgtgttcaa aaacgttata tttataggat aattatactc tatttctcaa
     5821 caagtaattg gttgtttggc cgagcggtct aaggcgcctg attcaagaaa tatcttgacc
     5881 gcagttaact gtgggaatac tcaggtatcg taagatgcaa gagttcgaat ctcttagcaa
     5941 ccattatttt tttcctcaac ataacgagaa cacacagggg cgctatcgca cagaatcaaa
     6001 ttcgatgact ggaaattttt tgttaatttc agaggtcgcc tgacgcatat acctttttca
     6061 actgaaaaat tgggagaaaa aggaaaggtg agagcgccgg aaccggcttt tcatatagaa
     6121 tagagaagcg ttcatgacta aatgcttgca tcacaatact tgaagttgac aatattattt
     6181 aaggacctat tgttttttcc aataggtggt tagcaatcgt cttactttct aacttttctt
     6241 accttttaca tttcagcaat atatatatat atatttcaag gatataccat tctaatgtct
     6301 gcccctaaga agatcgtcgt tttgccaggt gaccacgttg gtcaagaaat cacagccgaa
     6361 gccattaagg ttcttaaagc tatttctgat gttcgttcca atgtcaagtt cgatttcgaa
     6421 aatcatttaa ttggtggtgc tgctatcgat gctacaggtg ttccacttcc agatgaggcg
     6481 ctggaagcct ccaagaaggc tgatgccgtt ttgttaggtg ctgtgggtgg tcctaaatgg
     6541 ggtaccggta gtgttagacc tgaacaaggt ttactaaaaa tccgtaaaga acttcaattg
     6601 tacgccaact taagaccatg taactttgca tccgactctc ttttagactt atctccaatc
     6661 aagccacaat ttgctaaagg tactgacttc gttgttgtca gagaattagt gggaggtatt
     6721 tactttggta agagaaagga agacgatggt gatggtgtcg cttgggatag tgaacaatac
     6781 accgttccag aagtgcaaag aatcacaaga atggccgctt tcatggccct acaacatgag
     6841 ccaccattgc ctatttggtc cttggataaa gctaatgttt tggcctcttc aagattatgg
     6901 agaaaaactg tggaggaaac catcaagaac gaattcccta cattgaaggt tcaacatcaa
     6961 ttgattgatt ctgccgccat gatcctagtt aagaacccaa cccacctaaa tggtattata
     7021 atcaccagca acatgtttgg tgatatcatc tccgatgaag cctccgttat cccaggttcc
     7081 ttgggtttgt tgccatctgc gtccttggcc tctttgccag acaagaacac cgcatttggt
     7141 ttgtacgaac catgccacgg ttctgctcca gatttgccaa agaataaggt caaccctatc
     7201 gccactatct tgtctgctgc aatgatgttg aaattgtcat tgaacttgcc tgaagaaggt
     7261 aaggccattg aagatgcagt taaaaaggtt ttggatgcag gtatcagaac tggtgattta
     7321 ggtggttcca acagtaccac cgaagtcggt gatgctgtcg ccgaagaagt taagaaaatc
     7381 cttgcttaaa aagattctct ttttttatga tatttgtaca taaactttat aaatgaaatt
     7441 cataatagaa acgacacgaa attacaaaat ggaatatgtt catagggtag acgaaactat
     7501 atacgcaatc tacatacatt tatcaagaag gagaaaaagg aggatgtaaa ggaatacagg
     7561 taagcaaatt gatactaatg gctcaacgtg ataaggaaaa agaattgcac tttaacatta
     7621 atattgacaa ggaggagggc accacacaaa aagttaggtg taacagaaaa tcatgaaact
     7681 atgattccta atttatatat tggaggattt tctctaaaaa aaaaaaaata caacaaataa
     7741 aaaacactca atgacctgac catttgatgg agtttaagtc aataccttct tgaaccattt
     7801 cccataatgg tgaaagttcc ctcaagaatt ttactctgtc agaaacggcc ttaacgacgt
     7861 agtcgatatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc
     7921 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     7981 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     8041 cgaaacgcgc g
//
