LOCUS       sequence-232457-        6937 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6937)
  AUTHORS   Nathan DF, Lindquist S
  TITLE     Mutational analysis of Hsp90 function: interactions with a steroid 
            receptor and a protein kinase.
  JOURNAL   Mol Cell Biol 1995 Jul;15(7):3917-25.
  PUBMED    7791797
REFERENCE   2  (bases 1 to 6937)
  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: "Mol Cell 
            Biol"; date: "1995-07"; volume: "15(7)"; pages: "3917-25"
COMMENT     Sequence Label: pBK1
FEATURES             Location/Qualifiers
     source          1..6937
                     /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"
     misc_feature    22..78
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     CDS             complement(1395..2489)
                     /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"
     promoter        complement(2490..2895)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     promoter        complement(3822..3840)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3861..3877)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3861..3877)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3874..3896)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3885..3901
                     /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(3909..3915,3916..3933,3934..3939))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3954..3975
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4092..4109)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4263..4851)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4343..4362)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5022..5813,5814..5882))
                     /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     5645..5664
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5883..5987)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6055..6073
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6111..6133)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6233..6252
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(6319..6774)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6456..6477)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6668..6687
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6900..6922
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6914..6931
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6915..6931
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 taatacgact cactataggg cgaattcgag ctcggtaccc ggggatcctc tagagtcgac
       61 ctgcaggcat gcaagctttt aaccgtacta gatagtttat aacccattac gcatttgatt
      121 ataatttgct tcttaggcaa aattaatatt tacgttcttt tatattcttt ctttttgtat
      181 tcatagaaca gcagccatta ccaatagaaa gttaaaatag ccgccgatgc attttattac
      241 ccgcctttct gttttctggg cacttttctt tctagaaggt gaaagaacaa tttttctcgt
      301 tttctcgaac ttccaccaag cgttgggtaa tgagggagga gatttgtata aaaagagtgg
      361 catgtgaact gcctaccgta agtgacatga acacatgcat tatatttttt gtgatatatt
      421 ctttctcttg ttttcttttt cttgaaacgc tacagaacca atagaaaaat agaatcattc
      481 tgaaatatgg ctggtgaaac ttttgaattt caagctgaaa tcactcagtt gatgagtttg
      541 atctggtact ttggcccccc cattggtagc gggatttggt gaagctgcga gattgatgaa
      601 gaaagaattt gacaacgacc aagctcacat caaaagacta tccgataaat tagtcaaagg
      661 tctattatcc gctgaacata ccacgttgaa cggatctcca gatcatcgtt atccagggtg
      721 tgttaacgtt tctttcgcct acgtggaagg agaatcttta ttgatggcac taagggatat
      781 cgcattatcc tcgggttcag cctgtacatc tgcttcccta gaaccttctt atgttttaca
      841 tgcgctgggt aaggatgatg cattagccca ttcttccatc agatttggta ttggtagatt
      901 tagtactgaa gaggaggtcg actacgtcgt taaggccgtt tctgacagag taaaattctt
      961 gagggaactt tcaccattat gggaaatggt tcaagaaggt attgacttaa actccatcaa
     1021 atggtcaggt cattgagtgt tttttatttg ttgtattttt ttttttttag agaaaatcct
     1081 ccaatatata aattaggaat catagtttca tgattttctg ttacacctaa ctttttgtgt
     1141 ggtgccctcc tccttgtcaa tattaatgtt aaagtgcaat tctttttcct tatcacgttg
     1201 agccattagt atcaatttgc ttacctgtat tcctttacat cctccttttt ctccttcttg
     1261 ataaatgtat gtagattgcg tatatagttt cgtctaccct atgaacatat tccattttgt
     1321 aatttcgtgt cgtttctatt atgaatttca tttataaagt ttatgtacaa atatcataaa
     1381 aaaagagaat ctttttaagc aaggattttc ttaacttctt cggcgacagc atcaccgact
     1441 tcggtggtac tgttggaacc acctaaatca ccagttctga tacctgcatc caaaaccttt
     1501 ttaactgcat cttcaatggc cttaccttct tcaggcaagt tcaatgacaa tttcaacatc
     1561 attgcagcag acaagatagt ggcgataggg ttgaccttat tctttggcaa atctggagca
     1621 gaaccgtggc atggttcgta caaaccaaat gcggtgttct tgtctggcaa agaggccaag
     1681 gacgcagatg gcaacaaacc caaggaacct gggataacgg aggcttcatc ggagatgata
     1741 tcaccaaaca tgttgctggt gattataata ccatttaggt gggttgggtt cttaactagg
     1801 atcatggcgg cagaatcaat caattgatgt tgaaccttca atgtagggaa ttcgttcttg
     1861 atggtttcct ccacagtttt tctccataat cttgaagagg ccaaaacatt agctttatcc
     1921 aaggaccaaa taggcaatgg tggctcatgt tgtagggcca tgaaagcggc cattcttgtg
     1981 attctttgca cttctggaac ggtgtattgt tcactatccc aagcgacacc atcaccatcg
     2041 tcttcctttc tcttaccaaa gtaaatacct cccactaatt ctctgacaac aacgaagtca
     2101 gtacctttag caaattgtgg cttgattgga gataagtcta aaagagagtc ggatgcaaag
     2161 ttacatggtc ttaagttggc gtacaattga agttctttac ggatttttag taaaccttgt
     2221 tcaggtctaa cactaccggt accccattta ggaccaccca cagcacctaa caaaacggca
     2281 tcagccttct tggaggcttc cagcgcctca tctggaagtg gaacacctgt agcatcgata
     2341 gcagcaccac caattaaatg attttcgaaa tcgaacttga cattggaacg aacatcagaa
     2401 atagctttaa gaaccttaat ggcttcggct gtgatttctt gaccaacgtg gtcacctggc
     2461 aaaacgacga tcttcttagg ggcagacatt agaatggtat atccttgaaa tatatatata
     2521 tattgctgaa atgtaaaagg taagaaaagt tagaaagtaa gacgattgct aaccacctat
     2581 tggaaaaaac aataggtcct taaataatat tgtcaacttc aagtattgtg atgcaagcat
     2641 ttagtcatga acgcttctct attctatatg aaaagccggt tccggcgctc tcacctttcc
     2701 tttttctccc aatttttcag ttgaaaaagg tatatgcgtc aggcgacctc tgaaattaac
     2761 aaaaaatttc cagtcatcga atttgattct gtgcgatagc gcccctgtgt gttctcgtta
     2821 tgttgaggaa aaaaataatg gttgctaaga gattcgaact cttgcatctt acgatacctg
     2881 agtattccca cagttaactg cggtcaagat atttcttgaa tcaggcgcct tagaccgctc
     2941 ggccaaacaa ccaattactt gttgagaaat agagtataat tatcctataa atataacgtt
     3001 tttgaacaca catgaacaag gaagtacagg acaattgatt ttgaagagaa tgtggatttt
     3061 gatgtaattg ttgggattcc atttttaata aggcaataat attaggtatg tagatatact
     3121 agaagttctc ctcgaggatt taggaatcca taaaagggaa tctgcaattc tacacaattc
     3181 tataaatatt attatcatca ttttatatgt taatattcat tgatcctatt acattatcaa
     3241 tccttgcgtt tcagcttcca ctaatttaga tgactatttc tcatcatttg cgtcatcttc
     3301 taacaccgta tatgataata tactagtaac gtaaatacta gttagtagat gatagttgat
     3361 ttttattcca acataccacc cataatgtaa tagatccaag gccttgaagg atatcttggg
     3421 tgaccaagtg gagaaggttg ttgtttctta caaattgcta gatgctccag ctgccatcag
     3481 aactggtcaa ttcggctggt ctgctaacat ggaaagaatc atgaaggctc aagccttgag
     3541 agactcttcc atgtcctcct acatgtcttc caagaagact ttcgaaattt ctccaaaatc
     3601 tccaattatt aaggaattga aaaagagagt tgatgagggt ggtgcacaag ataagaccgt
     3661 caaagatttg actaacttat tattcgagac cgctttgttg acttctggtt tcagtttgga
     3721 agaaccaact tcttttgcat caagaataaa tagattgatt tctttaggtt tgaacattga
     3781 tgaggatgaa gaaacagaaa ccgctccaga agcttttgtt ccctttagtg agggttaatt
     3841 tcgagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     3901 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     3961 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     4021 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     4081 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     4141 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     4201 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     4261 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4321 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4381 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4441 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4501 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4561 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4621 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4681 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     4741 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     4801 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     4861 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     4921 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     4981 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     5041 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     5101 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     5161 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     5221 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5281 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5341 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5401 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5461 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5521 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5581 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5641 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     5701 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     5761 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     5821 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     5881 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     5941 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     6001 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     6061 cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     6121 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     6181 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca
     6241 gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg
     6301 agaaaatacc gcatcaggaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     6361 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     6421 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     6481 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     6541 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     6601 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     6661 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     6721 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtccattc
     6781 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     6841 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     6901 ccagtcacga cgttgtaaaa cgacggccag tgaattg
//
