LOCUS       sequence-241094-        7759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  2µm HIS3 CNB1mutEF3.
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: pTJK510
FEATURES             Location/Qualifiers
     source          1..7759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     52..68
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     52..68
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(1170..1186)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1171..1187)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(1215..1233)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1254..1270)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1254..1270)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1267..1289)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1278..1294
                     /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(1302..1308,1309..1326,1327..1332))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1347..1368
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1485..1502)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1656..2244)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1736..1755)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2415..3206,3207..3275))
                     /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     3038..3057
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3276..3380)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3965..5307
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(4928..4975)
                     /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     5687..5705
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5743..5765)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5865..5884
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        6031..6218
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             6219..6881
                     /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(7141..7596)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7278..7299)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7490..7509
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7722..7744
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7736..7753
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7737..7753
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 taatacgact cactataggg cgaattggag ctccaccgcg gtggcggccg ctctagaact
       61 agtggatcca cgctatgaaa aaggaaaaca aatgactctt ataatttgtt tccctcgact
      121 tctctattta aatccaagat tttaactaat aaactattta ataataaaac aattaactct
      181 ttatatggaa gccttggaaa ttagccgcca aaatgggata tacattccgt gcgaagtgac
      241 cgcgtggaag gccgggtatc atcttaaaaa tcactagttt cttttttagc ggaatgcaat
      301 aaaggtgctt tgtgctggtg tggtttacac ggaacatcta gtagctaaaa cttggtaact
      361 caatggtgat cagaatccat agaagcattt ttatttctta aaatgggtgc tgctccttcc
      421 aaaattgtgg atggtctttt agaagataca aattgtatgt acacttcgga gtgaggaaaa
      481 gaaagaaagg ggaaattaac cgattttact aacactgaca ctttgaacag ttgatagaga
      541 tgaaattgaa aggttaagga agagattcat gaaattagat agagatagct cagggtctat
      601 tgataaaaat gaatttatga gcattcctgg cgtttcgtca aaccctcttg ctggacgtat
      661 aatggaggtt ttcgatgctg ataatagtgg ggacgtggat tttcaagagt tcatcacagg
      721 attatccatt ttcagtgggc gtgggtccaa ggacgaaaag ttaagattcg ccttcaaaat
      781 ctacgccatt gacaaggacg gtttcatatc caatggtgag ttgttcatcg tgttgaagat
      841 tatggtaggt tctaatctgg acgatgaaca gctgcaacag atagtagata ggacgatagt
      901 ggaaaacgat agcgacggcg acggacgttt aagtttcgag gagtttaaga atgctatcga
      961 aaccacagaa gtggccaaga gtctgacatt gcaatacgat gtgtaagact aggggacact
     1021 tcattcattt atggtatgcc aatattttta agaaaagaag aataatacgc gatattgttt
     1081 tttaaggaag gaacgcacac tcgcccagtt agagtgctga tgatatatac atatatatat
     1141 gtatatgtaa caaacaataa gaagcttatc gataccgtcg acctcgaggg ggggcccggt
     1201 accagctttt gttcccttta gtgagggtta atttcgagct tggcgtaatc atggtcatag
     1261 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     1321 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     1381 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     1441 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     1501 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     1561 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     1621 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     1681 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     1741 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     1801 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     1861 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     1921 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     1981 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     2041 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     2101 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     2161 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     2221 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     2281 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     2341 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     2401 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     2461 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     2521 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     2581 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     2641 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     2701 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     2761 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     2821 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     2881 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     2941 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     3001 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     3061 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     3121 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     3181 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     3241 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     3301 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     3361 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctaattc cacggactat
     3421 agactatact agtatactcc gtctactgta cgatacactt ccgctcaggt ccttgtcctt
     3481 taacgaggcc ttaccactct tttgttactc tattgatcca gctcagcaaa ggcagtgtga
     3541 tctaagattc tatcttcgcg atgtagtaaa actagctaga ccgagaaaga gactagaaat
     3601 gcaaaaggca cttctacaat ggctgccatc attattatcc gatgtgacgc tgcagcttct
     3661 caatgatatt cgaatacgct ttgaggagat acagcctaat atccgacaaa ctgttttaca
     3721 gatttacgat cgtacttgtt acccatcatt gaattttgaa catccgaacc tgggagtttt
     3781 ccctgaaaca gatagtatat ttgaacctgt ataataatat atagtctagc gctttacgga
     3841 agacaatgta tgtatttcgg ttcctggaga aactattgca tctattgcat aggtaatctt
     3901 gcacgtcgca tccccggttc attttctgcg tttccatctt gcacttcaat agcatatctt
     3961 tgttaacgaa gcatctgtgc ttcattttgt agaacaaaaa tgcaacgcga gagcgctaat
     4021 ttttcaaaca aagaatctga gctgcatttt tacagaacag aaatgcaacg cgaaagcgct
     4081 attttaccaa cgaagaatct gtgcttcatt tttgtaaaac aaaaatgcaa cgcgagagcg
     4141 ctaatttttc aaacaaagaa tctgagctgc atttttacag aacagaaatg caacgcgaga
     4201 gcgctatttt accaacaaag aatctatact tcttttttgt tctacaaaaa tgcatcccga
     4261 gagcgctatt tttctaacaa agcatcttag attacttttt ttctcctttg tgcgctctat
     4321 aatgcagtct cttgataact ttttgcactg taggtccgtt aaggttagaa gaaggctact
     4381 ttggtgtcta ttttctcttc cataaaaaaa gcctgactcc acttcccgcg tttactgatt
     4441 actagcgaag ctgcgggtgc attttttcaa gataaaggca tccccgatta tattctatac
     4501 cgatgtggat tgcgcatact ttgtgaacag aaagtgatag cgttgatgat tcttcattgg
     4561 tcagaaaatt atgaacggtt tcttctattt tgtctctata tactacgtat aggaaatgtt
     4621 tacattttcg tattgttttc gattcactct atgaatagtt cttactacaa tttttttgtc
     4681 taaagagtaa tactagagat aaacataaaa aatgtagagg tcgagtttag atgcaagttc
     4741 aaggagcgaa aggtggatgg gtaggttata tagggatata gcacagagat atatagcaaa
     4801 gagatacttt tgagcaatgt ttgtggaagc ggtattcgca atattttagt agctcgttac
     4861 agtccggtgc gtttttggtt ttttgaaagt gcgtcttcag agcgcttttg gttttcaaaa
     4921 gcgctctgaa gttcctatac tttctagaga ataggaactt cggaatagga acttcaaagc
     4981 gtttccgaaa acgagcgctt ccgaaaatgc aacgcgagct gcgcacatac agctcactgt
     5041 tcacgtcgca cctatatctg cgtgttgcct gtatatatat atacatgaga agaacggcat
     5101 agtgcgtgtt tatgcttaaa tgcgtactta tatgcgtcta tttatgtagg atgaaaggta
     5161 gtctagtacc tcctgtgata ttatcccatt ccatgcgggg tatcgtatgc ttccttcagc
     5221 actacccttt agctgttcta tatgctgcca ctcctcaatt ggattagtct catccttcaa
     5281 tgctatcatt tcctttgata ttggatcata tgcatagtac cgagaaacta gtgcgaagta
     5341 gtgatcaggt attgctgtta tctgatgagt atacgttgtc ctggccacgg cagaagcacg
     5401 cttatcgctc caatttccca caacattagt caactccgtt aggcccttca ttgaaagaaa
     5461 tgaggtcatc aaatgtcttc caatgtgaga ttttgggcca ttttttatag caaagattga
     5521 ataaggcgca tttttcttca aagctttatt gtacgatctg actaagttat cttttaataa
     5581 ttggtattcc tgtttattgc ttgaagaatt gccggtccta tttactcgtt ttaggactgg
     5641 ttcagaattc taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac
     5701 gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     5761 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     5821 cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat
     5881 tgtactgaga gtgcaccata aattcccgtt ttaagagctt ggtgagcgct aggagtcact
     5941 gccaggtatc gtttgaacac ggcattagtc agggaagtca taacacagtc ctttcccgca
     6001 attttctttt tctattactc ttggcctcct ctagtacact ctatattttt ttatgcctcg
     6061 gtaatgattt tcattttttt ttttccacct agcggatgac tctttttttt tcttagcgat
     6121 tggcattatc acataatgaa ttatacatta tataaagtaa tgtgatttct tcgaagaata
     6181 tactaaaaaa tgagcaggca agataaacga aggcaaagat gacagagcag aaagccctag
     6241 taaagcgtat tacaaatgaa accaagattc agattgcgat ctctttaaag ggtggtcccc
     6301 tagcgataga gcactcgatc ttcccagaaa aagaggcaga agcagtagca gaacaggcca
     6361 cacaatcgca agtgattaac gtccacacag gtatagggtt tctggaccat atgatacatg
     6421 ctctggccaa gcattccggc tggtcgctaa tcgttgagtg cattggtgac ttacacatag
     6481 acgaccatca caccactgaa gactgcggga ttgctctcgg tcaagctttt aaagaggccc
     6541 taggggccgt gcgtggagta aaaaggtttg gatcaggatt tgcgcctttg gatgaggcac
     6601 tttccagagc ggtggtagat ctttcgaaca ggccgtacgc agttgtcgaa cttggtttgc
     6661 aaagggagaa agtaggagat ctctcttgcg agatgatccc gcattttctt gaaagctttg
     6721 cagaggctag cagaattacc ctccacgttg attgtctgcg aggcaagaat gatcatcacc
     6781 gtagtgagag tgcgttcaag gctcttgcgg ttgccataag agaagccacc tcgcccaatg
     6841 gtaccaacga tgttccctcc accaaaggtg ttcttatgta gtgacaccga ttatttaaag
     6901 ctgcagcata cgatatatat acatgtgtat atatgtatac ctatgaatgt cagtaagtat
     6961 gtatacgaac agtatgatac tgaagatgac aaggtaatgc atcattctat acgtgtcatt
     7021 ctgaacgagg cgcgctttcc ttttttcttt ttgctttttc tttttttttc tcttgaactc
     7081 gacggatcta tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
     7141 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     7201 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     7261 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     7321 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     7381 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     7441 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     7501 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     7561 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccat tcgccattca ggctgcgcaa
     7621 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg
     7681 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
     7741 aacgacggcc agtgaattg
//
