LOCUS       sequence-247765-        7334 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pHuji in yeast expression vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7334)
  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 7334)
  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: pKL08
FEATURES             Location/Qualifiers
     source          1..7334
                     /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      3963..4210
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3979..3997)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4228..4247)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4229..4247)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4257..4274)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4257..4273)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4266..4288)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4414..4869
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4501..4520)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4711..4732
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5170..5577
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             5578..6672
                     /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(7166..7185)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7285..7307
                     /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 gtttcgatgg tttctaaagg
     3241 tgaagagaac aacatggcta ttatcaaaga gttcatgagg tttaaggtcc acatggaagg
     3301 ttcagtgaac ggccatgagt tcgaaatcga gggtgaaggc gaaggtagac catacgaggc
     3361 gttccagact gccaaactga aagtcactaa aggtggacct ttaccgtttg cgtgggatat
     3421 cctttctcct caatttatgt atggttcaaa agtttatatt aaacatccag cagacattcc
     3481 tgattatttt aaattgtcat tccctgaagg ttttagatgg gagagagtca tgaatttcga
     3541 ggatggtggc ataatccatg ttaatcagga ttcttcttta caggatggtg tgttcattta
     3601 caaagttaag ttaaggggaa ctaactttcc ctcagatggt ccagtgatgc aaaagaagac
     3661 gatgggctgg gaagcttcag aagaaagaat gtacccagag gatggcgcct taaagtccga
     3721 aattaaatat cgtctgaaac tgaaagatgg cggccactac gcagctgaag taaagactac
     3781 ttataaagct aagaaaccag tacagttacc cggtgcctat attgtcgata taaaattgga
     3841 catcgtctct cataatgaag attatactat cgttgaacag tacgagaggg ctgagggcag
     3901 gcactctacc ggtggtatgg acgagttgta taaatgaagc ttatcgatac cgtcgactcg
     3961 agtcatgtaa ttagttatgt cacgcttaca ttcacgccct ccccccacat ccgctctaac
     4021 cgaaaaggaa ggagttagac aacctgaagt ctaggtccct atttattttt ttatagttat
     4081 gttagtatta agaacgttat ttatatttca aatttttctt ttttttctgt acagacgcgt
     4141 gtacgcatgt aacattatac tgaaaacctt gcttgagaag gttttgggac gctcgaaggc
     4201 tttaatttgc ggccggtacc caattcgccc tatagtgagt cgtattacgc gcgctcactg
     4261 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     4321 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     4381 tcccaacagt tgcgcagcct gaatggcgaa tggacgcgcc ctgtagcggc gcattaagcg
     4441 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     4501 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     4561 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     4621 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     4681 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     4741 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     4801 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     4861 ttacaatttc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     4921 atcgaccggt cgaggagaac ttctagtata tctacatacc taatattatt gccttattaa
     4981 aaatggaatc ccaacaatta catcaaaatc cacattctct tcaaaatcaa ttgtcctgta
     5041 cttccttgtt catgtgtgtt caaaaacgtt atatttatag gataattata ctctatttct
     5101 caacaagtaa ttggttgttt ggccgagcgg tctaaggcgc ctgattcaag aaatatcttg
     5161 accgcagtta actgtgggaa tactcaggta tcgtaagatg caagagttcg aatctcttag
     5221 caaccattat ttttttcctc aacataacga gaacacacag gggcgctatc gcacagaatc
     5281 aaattcgatg actggaaatt ttttgttaat ttcagaggtc gcctgacgca tatacctttt
     5341 tcaactgaaa aattgggaga aaaaggaaag gtgagagcgc cggaaccggc ttttcatata
     5401 gaatagagaa gcgttcatga ctaaatgctt gcatcacaat acttgaagtt gacaatatta
     5461 tttaaggacc tattgttttt tccaataggt ggttagcaat cgtcttactt tctaactttt
     5521 cttacctttt acatttcagc aatatatata tatatatttc aaggatatac cattctaatg
     5581 tctgccccta agaagatcgt cgttttgcca ggtgaccacg ttggtcaaga aatcacagcc
     5641 gaagccatta aggttcttaa agctatttct gatgttcgtt ccaatgtcaa gttcgatttc
     5701 gaaaatcatt taattggtgg tgctgctatc gatgctacag gtgttccact tccagatgag
     5761 gcgctggaag cctccaagaa ggctgatgcc gttttgttag gtgctgtggg tggtcctaaa
     5821 tggggtaccg gtagtgttag acctgaacaa ggtttactaa aaatccgtaa agaacttcaa
     5881 ttgtacgcca acttaagacc atgtaacttt gcatccgact ctcttttaga cttatctcca
     5941 atcaagccac aatttgctaa aggtactgac ttcgttgttg tcagagaatt agtgggaggt
     6001 atttactttg gtaagagaaa ggaagacgat ggtgatggtg tcgcttggga tagtgaacaa
     6061 tacaccgttc cagaagtgca aagaatcaca agaatggccg ctttcatggc cctacaacat
     6121 gagccaccat tgcctatttg gtccttggat aaagctaatg ttttggcctc ttcaagatta
     6181 tggagaaaaa ctgtggagga aaccatcaag aacgaattcc ctacattgaa ggttcaacat
     6241 caattgattg attctgccgc catgatccta gttaagaacc caacccacct aaatggtatt
     6301 ataatcacca gcaacatgtt tggtgatatc atctccgatg aagcctccgt tatcccaggt
     6361 tccttgggtt tgttgccatc tgcgtccttg gcctctttgc cagacaagaa caccgcattt
     6421 ggtttgtacg aaccatgcca cggttctgct ccagatttgc caaagaataa ggtcaaccct
     6481 atcgccacta tcttgtctgc tgcaatgatg ttgaaattgt cattgaactt gcctgaagaa
     6541 ggtaaggcca ttgaagatgc agttaaaaag gttttggatg caggtatcag aactggtgat
     6601 ttaggtggtt ccaacagtac caccgaagtc ggtgatgctg tcgccgaaga agttaagaaa
     6661 atccttgctt aaaaagattc tcttttttta tgatatttgt acataaactt tataaatgaa
     6721 attcataata gaaacgacac gaaattacaa aatggaatat gttcataggg tagacgaaac
     6781 tatatacgca atctacatac atttatcaag aaggagaaaa aggaggatgt aaaggaatac
     6841 aggtaagcaa attgatacta atggctcaac gtgataagga aaaagaattg cactttaaca
     6901 ttaatattga caaggaggag ggcaccacac aaaaagttag gtgtaacaga aaatcatgaa
     6961 actatgattc ctaatttata tattggagga ttttctctaa aaaaaaaaaa atacaacaaa
     7021 taaaaaacac tcaatgacct gaccatttga tggagtttaa gtcaatacct tcttgaacca
     7081 tttcccataa tggtgaaagt tccctcaaga attttactct gtcagaaacg gccttaacga
     7141 cgtagtcgat atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     7201 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     7261 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     7321 caccgaaacg cgcg
//
