LOCUS       sequence-238887-        7759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  2µm HIS3 CNB1mutEF1.
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: pTJK508
FEATURES             Location/Qualifiers
     source          1..7759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(602..618)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(603..619)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(647..665)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(686..702)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(686..702)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(699..721)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    710..726
                     /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(734..740,741..758,759..764))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    779..800
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(917..934)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1088..1676)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1168..1187)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1847..2638,2639..2707))
                     /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     2470..2489
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2708..2812)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3397..4739
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(4360..4407)
                     /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     5119..5137
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5175..5197)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5297..5316
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5463..5650
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             5651..6313
                     /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(6573..7028)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6710..6731)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6922..6941
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7154..7176
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7168..7185
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7169..7185
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7192..7211
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7192..7210
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     7243..7259
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7243..7259
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
ORIGIN
        1 atgaaattag ctagagatag ctcagggtct attgataaaa atgaatttat gagcattcct
       61 ggcgtttcgt caaaccctct tgctggacgt ataatggagg ttttcgatgc tgataatagt
      121 ggggacgtgg attttcaaga gttcatcaca ggattatcca ttttcagtgg gcgtgggtcc
      181 aaggacgaaa agttaagatt cgccttcaaa atctacgaca ttgacaagga cggtttcata
      241 tccaatggtg agttgttcat cgtgttgaag attatggtag gttctaatct ggacgatgaa
      301 cagctgcaac agatagtaga taggacgata gtggaaaacg atagcgacgg cgacggacgt
      361 ttaagtttcg aggagtttaa gaatgctatc gaaaccacag aagtggccaa gagtctgaca
      421 ttgcaatacg atgtgtaaga ctaggggaca cttcattcat ttatggtatg ccaatatttt
      481 taagaaaaga agaataatac gcgatattgt tttttaagga aggaacgcac actcgcccag
      541 ttagagtgct gatgatatat acatatatat atgtatatgt aacaaacaat aagaagctta
      601 tcgataccgt cgacctcgag ggggggcccg gtaccagctt ttgttccctt tagtgagggt
      661 taatttcgag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
      721 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
      781 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
      841 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
      901 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
      961 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     1021 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     1081 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     1141 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     1201 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     1261 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     1321 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     1381 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     1441 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     1501 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     1561 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     1621 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     1681 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     1741 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     1801 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     1861 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     1921 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     1981 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     2041 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     2101 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     2161 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     2221 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     2281 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     2341 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     2401 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     2461 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     2521 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     2581 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     2641 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     2701 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     2761 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     2821 cccgaaaagt gccacctaat tccacggact atagactata ctagtatact ccgtctactg
     2881 tacgatacac ttccgctcag gtccttgtcc tttaacgagg ccttaccact cttttgttac
     2941 tctattgatc cagctcagca aaggcagtgt gatctaagat tctatcttcg cgatgtagta
     3001 aaactagcta gaccgagaaa gagactagaa atgcaaaagg cacttctaca atggctgcca
     3061 tcattattat ccgatgtgac gctgcagctt ctcaatgata ttcgaatacg ctttgaggag
     3121 atacagccta atatccgaca aactgtttta cagatttacg atcgtacttg ttacccatca
     3181 ttgaattttg aacatccgaa cctgggagtt ttccctgaaa cagatagtat atttgaacct
     3241 gtataataat atatagtcta gcgctttacg gaagacaatg tatgtatttc ggttcctgga
     3301 gaaactattg catctattgc ataggtaatc ttgcacgtcg catccccggt tcattttctg
     3361 cgtttccatc ttgcacttca atagcatatc tttgttaacg aagcatctgt gcttcatttt
     3421 gtagaacaaa aatgcaacgc gagagcgcta atttttcaaa caaagaatct gagctgcatt
     3481 tttacagaac agaaatgcaa cgcgaaagcg ctattttacc aacgaagaat ctgtgcttca
     3541 tttttgtaaa acaaaaatgc aacgcgagag cgctaatttt tcaaacaaag aatctgagct
     3601 gcatttttac agaacagaaa tgcaacgcga gagcgctatt ttaccaacaa agaatctata
     3661 cttctttttt gttctacaaa aatgcatccc gagagcgcta tttttctaac aaagcatctt
     3721 agattacttt ttttctcctt tgtgcgctct ataatgcagt ctcttgataa ctttttgcac
     3781 tgtaggtccg ttaaggttag aagaaggcta ctttggtgtc tattttctct tccataaaaa
     3841 aagcctgact ccacttcccg cgtttactga ttactagcga agctgcgggt gcattttttc
     3901 aagataaagg catccccgat tatattctat accgatgtgg attgcgcata ctttgtgaac
     3961 agaaagtgat agcgttgatg attcttcatt ggtcagaaaa ttatgaacgg tttcttctat
     4021 tttgtctcta tatactacgt ataggaaatg tttacatttt cgtattgttt tcgattcact
     4081 ctatgaatag ttcttactac aatttttttg tctaaagagt aatactagag ataaacataa
     4141 aaaatgtaga ggtcgagttt agatgcaagt tcaaggagcg aaaggtggat gggtaggtta
     4201 tatagggata tagcacagag atatatagca aagagatact tttgagcaat gtttgtggaa
     4261 gcggtattcg caatatttta gtagctcgtt acagtccggt gcgtttttgg ttttttgaaa
     4321 gtgcgtcttc agagcgcttt tggttttcaa aagcgctctg aagttcctat actttctaga
     4381 gaataggaac ttcggaatag gaacttcaaa gcgtttccga aaacgagcgc ttccgaaaat
     4441 gcaacgcgag ctgcgcacat acagctcact gttcacgtcg cacctatatc tgcgtgttgc
     4501 ctgtatatat atatacatga gaagaacggc atagtgcgtg tttatgctta aatgcgtact
     4561 tatatgcgtc tatttatgta ggatgaaagg tagtctagta cctcctgtga tattatccca
     4621 ttccatgcgg ggtatcgtat gcttccttca gcactaccct ttagctgttc tatatgctgc
     4681 cactcctcaa ttggattagt ctcatccttc aatgctatca tttcctttga tattggatca
     4741 tatgcatagt accgagaaac tagtgcgaag tagtgatcag gtattgctgt tatctgatga
     4801 gtatacgttg tcctggccac ggcagaagca cgcttatcgc tccaatttcc cacaacatta
     4861 gtcaactccg ttaggccctt cattgaaaga aatgaggtca tcaaatgtct tccaatgtga
     4921 gattttgggc cattttttat agcaaagatt gaataaggcg catttttctt caaagcttta
     4981 ttgtacgatc tgactaagtt atcttttaat aattggtatt cctgtttatt gcttgaagaa
     5041 ttgccggtcc tatttactcg ttttaggact ggttcagaat tctaagaaac cattattatc
     5101 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt
     5161 gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa
     5221 gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg
     5281 ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca taaattcccg
     5341 ttttaagagc ttggtgagcg ctaggagtca ctgccaggta tcgtttgaac acggcattag
     5401 tcagggaagt cataacacag tcctttcccg caattttctt tttctattac tcttggcctc
     5461 ctctagtaca ctctatattt ttttatgcct cggtaatgat tttcattttt ttttttccac
     5521 ctagcggatg actctttttt tttcttagcg attggcatta tcacataatg aattatacat
     5581 tatataaagt aatgtgattt cttcgaagaa tatactaaaa aatgagcagg caagataaac
     5641 gaaggcaaag atgacagagc agaaagccct agtaaagcgt attacaaatg aaaccaagat
     5701 tcagattgcg atctctttaa agggtggtcc cctagcgata gagcactcga tcttcccaga
     5761 aaaagaggca gaagcagtag cagaacaggc cacacaatcg caagtgatta acgtccacac
     5821 aggtataggg tttctggacc atatgataca tgctctggcc aagcattccg gctggtcgct
     5881 aatcgttgag tgcattggtg acttacacat agacgaccat cacaccactg aagactgcgg
     5941 gattgctctc ggtcaagctt ttaaagaggc cctaggggcc gtgcgtggag taaaaaggtt
     6001 tggatcagga tttgcgcctt tggatgaggc actttccaga gcggtggtag atctttcgaa
     6061 caggccgtac gcagttgtcg aacttggttt gcaaagggag aaagtaggag atctctcttg
     6121 cgagatgatc ccgcattttc ttgaaagctt tgcagaggct agcagaatta ccctccacgt
     6181 tgattgtctg cgaggcaaga atgatcatca ccgtagtgag agtgcgttca aggctcttgc
     6241 ggttgccata agagaagcca cctcgcccaa tggtaccaac gatgttccct ccaccaaagg
     6301 tgttcttatg tagtgacacc gattatttaa agctgcagca tacgatatat atacatgtgt
     6361 atatatgtat acctatgaat gtcagtaagt atgtatacga acagtatgat actgaagatg
     6421 acaaggtaat gcatcattct atacgtgtca ttctgaacga ggcgcgcttt ccttttttct
     6481 ttttgctttt tctttttttt tctcttgaac tcgacggatc tatgcggtgt gaaataccgc
     6541 acagatgcgt aaggagaaaa taccgcatca ggaaattgta agcgttaata ttttgttaaa
     6601 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     6661 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     6721 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     6781 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     6841 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     6901 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     6961 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     7021 gggcgcgtcc attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc
     7081 tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta
     7141 acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt gtaatacgac
     7201 tcactatagg gcgaattgga gctccaccgc ggtggcggcc gctctagaac tagtggatcc
     7261 acgctatgaa aaaggaaaac aaatgactct tataatttgt ttccctcgac ttctctattt
     7321 aaatccaaga ttttaactaa taaactattt aataataaaa caattaactc tttatatgga
     7381 agccttggaa attagccgcc aaaatgggat atacattccg tgcgaagtga ccgcgtggaa
     7441 ggccgggtat catcttaaaa atcactagtt tcttttttag cggaatgcaa taaaggtgct
     7501 ttgtgctggt gtggtttaca cggaacatct agtagctaaa acttggtaac tcaatggtga
     7561 tcagaatcca tagaagcatt tttatttctt aaaatgggtg ctgctccttc caaaattgtg
     7621 gatggtcttt tagaagatac aaattgtatg tacacttcgg agtgaggaaa agaaagaaag
     7681 gggaaattaa ccgattttac taacactgac actttgaaca gttgatagag atgaaattga
     7741 aaggttaagg aagagattc
//
