LOCUS       sequence-497704-        5081 bp    DNA     circular SYN 16-JAN-2026
DEFINITION  expresses yeast Cak1 in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5081)
  AUTHORS   Rimal A, Swayne TM, Kamdar ZP, Tewey MA, Winter E
  TITLE     Isc10, an inhibitor of the Smk1 MAPK, prevents activation loop 
            autophosphorylation and substrate phosphorylation through separate 
            mechanisms.
  JOURNAL   J Biol Chem. 2022 Oct;298(10):102450. doi: 
            10.1016/j.jbc.2022.102450. Epub 2022 Sep 3.
  PUBMED    36063999
REFERENCE   2  (bases 1 to 5081)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 16, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Biol 
            Chem."; date: "2022-10"; volume: "298(10):102450. doi"; pages: " 
            10.1016/j.jbc.2022.102450. Epub 2022 Sep 3"
COMMENT     Sequence Label: pJTB122
FEATURES             Location/Qualifiers
     source          1..5081
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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)."
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        214..232
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    233..257
                     /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)."
     CDS             1437..1481
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1519..1537)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1533..1580
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1801..2460)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2375..2394
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2461..2563)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(2955..2972)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3089..3634)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(3169..3188)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3859..4941)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     4903..4922
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4942..5019)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(5065..5081,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cgaattcgag
      121 ctcggcgcgc ctgcaggtcg acaagcttgc ggccgcataa tgcttaagtc gaacagaaag
      181 taatcgtatt gtacacggcc gcataatcga aattaatacg actcactata ggggaattgt
      241 gagcggataa caattcccca tcttagtata ttagttaagt ataagaagga gatatacata
      301 tggcagatct catgaaactg gatagtatag acattacaca ctgtcagttg gtcaaatcta
      361 ctagaactgc taggatttat aggtcggata catatgccat taaatgtcta gcactagatt
      421 tcgatatccc gccacataac gccaaattcg aagtatcgat attaaacaaa ctgggcaaca
      481 aatgtaagca catcttacct cttctagagt ctaaggctac cgataataat gacctattgt
      541 tgctgtttcc ctttgaagag atgaaccttt atgagttcat gcaaatgcac tataaaagag
      601 atagaagaaa aaaaaatccc tattacgatt tgctaaatcc cagtatccca attgttgcgg
      661 accccccagt tcagaaatat actaatcaat tggacgtcaa tcggtattct ttgtcctttt
      721 tctggcaaat ggttgaaggg attgcattct tacatgagaa caagatcatt caccgcgaca
      781 tcaaaccgca aaatatcatg ctaacgaaca ataccagcac cgtatcccca aagttgtaca
      841 taattgattt tggcatctca tatgacatgg caaataactc acaaacaagt gcggaaccca
      901 tggacagcaa ggtgacggat ataagcacag gaatttacaa ggccccagaa gtgctttttg
      961 gagtaaaatg ctatgatggt ggcgtggacg tgtggtcgtt gttgataatt atttctcagt
     1021 ggttccagag agaaacaagc cgtatggggc acgttccggc catgattgat gacggcagcg
     1081 acgacatgaa ctcagatgga agcgatttca gactgatttg ctcaatattc gaaaagttgg
     1141 gcataccgtc cattcagaaa tgggaggagg ttgcgcaaca cggctcggtt gatgcatttg
     1201 ttggtatgtt tggtgcagat ggcgatggta agtatgtact ggaccaggag aaagatgtac
     1261 agattagcat cgttgagagg aatatgcctc gactggacga gattgcggat gtcaaagtca
     1321 agcagaagtt cattaattgt atcctgggga tggtttcatt ttcaccaaac gaaagatgga
     1381 gctgtcaaag aatcttgcaa gaactagaaa agccataagg taccctcgag tctggtaaag
     1441 aaaccgctgc tgcgaaattt gaacgccagc acatggactc gtctactagc gcagcttaat
     1501 taacctaggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     1561 gggtcttgag gggttttttg ctgaaacctc aggcatttga gaagcacacg gtcacactgc
     1621 ttccggtagt caataaaccg gtaaaccagc aatagacata agcggctatt taacgaccct
     1681 gccctgaacc gacgaccggg tcgaatttgc tttcgaattt ctgccattca tccgcttatt
     1741 atcacttatt caggcgtagc aaccaggcgt ttaagggcac caataactgc cttaaaaaaa
     1801 ttacgccccg ccctgccact catcgcagta ctgttgtaat tcattaagca ttctgccgac
     1861 atggaagcca tcacaaacgg catgatgaac ctgaatcgcc agcggcatca gcaccttgtc
     1921 gccttgcgta taatatttgc ccatagtgaa aacgggggcg aagaagttgt ccatattggc
     1981 cacgtttaaa tcaaaactgg tgaaactcac ccagggattg gctgagacga aaaacatatt
     2041 ctcaataaac cctttaggga aataggccag gttttcaccg taacacgcca catcttgcga
     2101 atatatgtgt agaaactgcc ggaaatcgtc gtggtattca ctccagagcg atgaaaacgt
     2161 ttcagtttgc tcatggaaaa cggtgtaaca agggtgaaca ctatcccata tcaccagctc
     2221 accgtctttc attgccatac ggaactccgg atgagcattc atcaggcggg caagaatgtg
     2281 aataaaggcc ggataaaact tgtgcttatt tttctttacg gtctttaaaa aggccgtaat
     2341 atccagctga acggtctggt tataggtaca ttgagcaact gactgaaatg cctcaaaatg
     2401 ttctttacga tgccattggg atatatcaac ggtggtatat ccagtgattt ttttctccat
     2461 tttagcttcc ttagctcctg aaaatctcga taactcaaaa aatacgcccg gtagtgatct
     2521 tatttcatta tggtgaaagt tggaacctct tacgtgccga tcaacgtctc attttcgcca
     2581 aaagttggcc cagggcttcc cggtatcaac agggacacca ggatttattt attctgcgaa
     2641 gtgatcttcc gtcacaggta tttattcggc gcaaagtgcg tcgggtgatg ctgccaactt
     2701 actgatttag tgtatgatgg tgtttttgag gtgctccagt ggcttctgtt tctatcagct
     2761 gtccctcctg ttcagctact gacggggtgg tgcgtaacgg caaaagcacc gccggacatc
     2821 agcgctagcg gagtgtatac tggcttacta tgttggcact gatgagggtg tcagtgaagt
     2881 gcttcatgtg gcaggagaaa aaaggctgca ccggtgcgtc agcagaatat gtgatacagg
     2941 atatattccg cttcctcgct cactgactcg ctacgctcgg tcgttcgact gcggcgagcg
     3001 gaaatggctt acgaacgggg cggagatttc ctggaagatg ccaggaagat acttaacagg
     3061 gaagtgagag ggccgcggca aagccgtttt tccataggct ccgcccccct gacaagcatc
     3121 acgaaatctg acgctcaaat cagtggtggc gaaacccgac aggactataa agataccagg
     3181 cgtttccccc tggcggctcc ctcgtgcgct ctcctgttcc tgcctttcgg tttaccggtg
     3241 tcattccgct gttatggccg cgtttgtctc attccacgcc tgacactcag ttccgggtag
     3301 gcagttcgct ccaagctgga ctgtatgcac gaaccccccg ttcagtccga ccgctgcgcc
     3361 ttatccggta actatcgtct tgagtccaac ccggaaagac atgcaaaagc accactggca
     3421 gcagccactg gtaattgatt tagaggagtt agtcttgaag tcatgcgccg gttaaggcta
     3481 aactgaaagg acaagttttg gtgactgcgc tcctccaagc cagttacctc ggttcaaaga
     3541 gttggtagct cagagaacct tcgaaaaacc gccctgcaag gcggtttttt cgttttcaga
     3601 gcaagagatt acgcgcagac caaaacgatc tcaagaagat catcttatta atcagataaa
     3661 atatttctag atttcagtgc aatttatctc ttcaaatgta gcacctgaag tcagccccat
     3721 acgatataag ttgtaattct catgttagtc atgccccgcg cccaccggaa ggagctgact
     3781 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
     3841 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3901 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
     3961 ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
     4021 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
     4081 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
     4141 gtccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
     4201 atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg
     4261 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
     4321 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
     4381 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
     4441 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     4501 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
     4561 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     4621 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     4681 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     4741 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     4801 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     4861 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     4921 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
     4981 tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct
     5041 tatgcgactc ctgcattagg aaattaatac gactcactat a
//
