LOCUS       sequence-239345-        7759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  2µm HIS3 CNB1mutEF2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7759)
  AUTHORS   Connolly S, Quasi-Woode D, Waldron L, Eberly C, Waters K, Muller EM,
            Kingsbury TJ
  TITLE     Calcineurin Regulatory Subunit Calcium-Binding Domains 
            Differentially Contribute to Calcineurin Signaling in Saccharomyces 
            cerevisiae.
  JOURNAL   Genetics. 2018 Jul;209(3):801-813. doi: 10.1534/genetics.118.300911.
            Epub 2018 May 7.
  PUBMED    29735720
REFERENCE   2  (bases 1 to 7759)
  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: 
            "10.1534/genetics.118.300911"; journalName: "Genetics"; date: 
            "2018-07"; volume: "209"; issue: "3"; pages: "801-813"
COMMENT     Sequence Label: pTJK509
FEATURES             Location/Qualifiers
     source          1..7759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1..17
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(1119..1135)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1120..1136)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(1164..1182)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1203..1219)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1203..1219)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1216..1238)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1227..1243
                     /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(1251..1257,1258..1275,1276..1281))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1296..1317
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1434..1451)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1605..2193)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1685..1704)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2364..3155,3156..3224))
                     /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     2987..3006
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3225..3329)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3914..5256
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(4877..4924)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     5636..5654
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5692..5714)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5814..5833
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5980..6167
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             6168..6830
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(7090..7545)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7227..7248)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7439..7458
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7671..7693
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7685..7702
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7686..7702
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7709..7728
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7709..7727
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 tctagaacta gtggatccac gctatgaaaa aggaaaacaa atgactctta taatttgttt
       61 ccctcgactt ctctatttaa atccaagatt ttaactaata aactatttaa taataaaaca
      121 attaactctt tatatggaag ccttggaaat tagccgccaa aatgggatat acattccgtg
      181 cgaagtgacc gcgtggaagg ccgggtatca tcttaaaaat cactagtttc ttttttagcg
      241 gaatgcaata aaggtgcttt gtgctggtgt ggtttacacg gaacatctag tagctaaaac
      301 ttggtaactc aatggtgatc agaatccata gaagcatttt tatttcttaa aatgggtgct
      361 gctccttcca aaattgtgga tggtctttta gaagatacaa attgtatgta cacttcggag
      421 tgaggaaaag aaagaaaggg gaaattaacc gattttacta acactgacac tttgaacagt
      481 tgatagagat gaaattgaaa ggttaaggaa gagattcatg aaattagata gagatagctc
      541 agggtctatt gataaaaatg aatttatgag cattcctggc gtttcgtcaa accctcttgc
      601 tggacgtata atggaggttt tcgctgctga taatagtggg gacgtggatt ttcaagagtt
      661 catcacagga ttatccattt tcagtgggcg tgggtccaag gacgaaaagt taagattcgc
      721 cttcaaaatc tacgacattg acaaggacgg tttcatatcc aatggtgagt tgttcatcgt
      781 gttgaagatt atggtaggtt ctaatctgga cgatgaacag ctgcaacaga tagtagatag
      841 gacgatagtg gaaaacgata gcgacggcga cggacgttta agtttcgagg agtttaagaa
      901 tgctatcgaa accacagaag tggccaagag tctgacattg caatacgatg tgtaagacta
      961 ggggacactt cattcattta tggtatgcca atatttttaa gaaaagaaga ataatacgcg
     1021 atattgtttt ttaaggaagg aacgcacact cgcccagtta gagtgctgat gatatataca
     1081 tatatatatg tatatgtaac aaacaataag aagcttatcg ataccgtcga cctcgagggg
     1141 gggcccggta ccagcttttg ttccctttag tgagggttaa tttcgagctt ggcgtaatca
     1201 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     1261 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     1321 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     1381 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     1441 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     1501 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     1561 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     1621 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     1681 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     1741 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     1801 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     1861 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     1921 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     1981 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2041 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2101 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     2161 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     2221 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     2281 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     2341 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     2401 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     2461 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     2521 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     2581 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     2641 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     2701 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     2761 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     2821 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     2881 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     2941 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     3001 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     3061 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     3121 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     3181 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     3241 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     3301 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaattcc
     3361 acggactata gactatacta gtatactccg tctactgtac gatacacttc cgctcaggtc
     3421 cttgtccttt aacgaggcct taccactctt ttgttactct attgatccag ctcagcaaag
     3481 gcagtgtgat ctaagattct atcttcgcga tgtagtaaaa ctagctagac cgagaaagag
     3541 actagaaatg caaaaggcac ttctacaatg gctgccatca ttattatccg atgtgacgct
     3601 gcagcttctc aatgatattc gaatacgctt tgaggagata cagcctaata tccgacaaac
     3661 tgttttacag atttacgatc gtacttgtta cccatcattg aattttgaac atccgaacct
     3721 gggagttttc cctgaaacag atagtatatt tgaacctgta taataatata tagtctagcg
     3781 ctttacggaa gacaatgtat gtatttcggt tcctggagaa actattgcat ctattgcata
     3841 ggtaatcttg cacgtcgcat ccccggttca ttttctgcgt ttccatcttg cacttcaata
     3901 gcatatcttt gttaacgaag catctgtgct tcattttgta gaacaaaaat gcaacgcgag
     3961 agcgctaatt tttcaaacaa agaatctgag ctgcattttt acagaacaga aatgcaacgc
     4021 gaaagcgcta ttttaccaac gaagaatctg tgcttcattt ttgtaaaaca aaaatgcaac
     4081 gcgagagcgc taatttttca aacaaagaat ctgagctgca tttttacaga acagaaatgc
     4141 aacgcgagag cgctatttta ccaacaaaga atctatactt cttttttgtt ctacaaaaat
     4201 gcatcccgag agcgctattt ttctaacaaa gcatcttaga ttactttttt tctcctttgt
     4261 gcgctctata atgcagtctc ttgataactt tttgcactgt aggtccgtta aggttagaag
     4321 aaggctactt tggtgtctat tttctcttcc ataaaaaaag cctgactcca cttcccgcgt
     4381 ttactgatta ctagcgaagc tgcgggtgca ttttttcaag ataaaggcat ccccgattat
     4441 attctatacc gatgtggatt gcgcatactt tgtgaacaga aagtgatagc gttgatgatt
     4501 cttcattggt cagaaaatta tgaacggttt cttctatttt gtctctatat actacgtata
     4561 ggaaatgttt acattttcgt attgttttcg attcactcta tgaatagttc ttactacaat
     4621 ttttttgtct aaagagtaat actagagata aacataaaaa atgtagaggt cgagtttaga
     4681 tgcaagttca aggagcgaaa ggtggatggg taggttatat agggatatag cacagagata
     4741 tatagcaaag agatactttt gagcaatgtt tgtggaagcg gtattcgcaa tattttagta
     4801 gctcgttaca gtccggtgcg tttttggttt tttgaaagtg cgtcttcaga gcgcttttgg
     4861 ttttcaaaag cgctctgaag ttcctatact ttctagagaa taggaacttc ggaataggaa
     4921 cttcaaagcg tttccgaaaa cgagcgcttc cgaaaatgca acgcgagctg cgcacataca
     4981 gctcactgtt cacgtcgcac ctatatctgc gtgttgcctg tatatatata tacatgagaa
     5041 gaacggcata gtgcgtgttt atgcttaaat gcgtacttat atgcgtctat ttatgtagga
     5101 tgaaaggtag tctagtacct cctgtgatat tatcccattc catgcggggt atcgtatgct
     5161 tccttcagca ctacccttta gctgttctat atgctgccac tcctcaattg gattagtctc
     5221 atccttcaat gctatcattt cctttgatat tggatcatat gcatagtacc gagaaactag
     5281 tgcgaagtag tgatcaggta ttgctgttat ctgatgagta tacgttgtcc tggccacggc
     5341 agaagcacgc ttatcgctcc aatttcccac aacattagtc aactccgtta ggcccttcat
     5401 tgaaagaaat gaggtcatca aatgtcttcc aatgtgagat tttgggccat tttttatagc
     5461 aaagattgaa taaggcgcat ttttcttcaa agctttattg tacgatctga ctaagttatc
     5521 ttttaataat tggtattcct gtttattgct tgaagaattg ccggtcctat ttactcgttt
     5581 taggactggt tcagaattct aagaaaccat tattatcatg acattaacct ataaaaatag
     5641 gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     5701 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     5761 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc
     5821 agagcagatt gtactgagag tgcaccataa attcccgttt taagagcttg gtgagcgcta
     5881 ggagtcactg ccaggtatcg tttgaacacg gcattagtca gggaagtcat aacacagtcc
     5941 tttcccgcaa ttttcttttt ctattactct tggcctcctc tagtacactc tatatttttt
     6001 tatgcctcgg taatgatttt catttttttt tttccaccta gcggatgact cttttttttt
     6061 cttagcgatt ggcattatca cataatgaat tatacattat ataaagtaat gtgatttctt
     6121 cgaagaatat actaaaaaat gagcaggcaa gataaacgaa ggcaaagatg acagagcaga
     6181 aagccctagt aaagcgtatt acaaatgaaa ccaagattca gattgcgatc tctttaaagg
     6241 gtggtcccct agcgatagag cactcgatct tcccagaaaa agaggcagaa gcagtagcag
     6301 aacaggccac acaatcgcaa gtgattaacg tccacacagg tatagggttt ctggaccata
     6361 tgatacatgc tctggccaag cattccggct ggtcgctaat cgttgagtgc attggtgact
     6421 tacacataga cgaccatcac accactgaag actgcgggat tgctctcggt caagctttta
     6481 aagaggccct aggggccgtg cgtggagtaa aaaggtttgg atcaggattt gcgcctttgg
     6541 atgaggcact ttccagagcg gtggtagatc tttcgaacag gccgtacgca gttgtcgaac
     6601 ttggtttgca aagggagaaa gtaggagatc tctcttgcga gatgatcccg cattttcttg
     6661 aaagctttgc agaggctagc agaattaccc tccacgttga ttgtctgcga ggcaagaatg
     6721 atcatcaccg tagtgagagt gcgttcaagg ctcttgcggt tgccataaga gaagccacct
     6781 cgcccaatgg taccaacgat gttccctcca ccaaaggtgt tcttatgtag tgacaccgat
     6841 tatttaaagc tgcagcatac gatatatata catgtgtata tatgtatacc tatgaatgtc
     6901 agtaagtatg tatacgaaca gtatgatact gaagatgaca aggtaatgca tcattctata
     6961 cgtgtcattc tgaacgaggc gcgctttcct tttttctttt tgctttttct ttttttttct
     7021 cttgaactcg acggatctat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     7081 cgcatcagga aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     7141 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     7201 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     7261 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     7321 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     7381 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     7441 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     7501 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtccatt cgccattcag
     7561 gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac gccagctggc
     7621 gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg
     7681 acgttgtaaa acgacggcca gtgaattgta atacgactca ctatagggcg aattggagct
     7741 ccaccgcggt ggcggccgc
//
