LOCUS       sequence-237469-        7575 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a truncated construct of histone deacetylase 8 for 
            crystallography bearing an N-terminal tobacco etch virus 
            protease-cleavable hexahistidine-maltose binding protein tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7575)
  AUTHORS   Porter NJ, Christianson DW
  TITLE     Binding of the Microbial Cyclic Tetrapeptide Trapoxin A to the Class
            I Histone Deacetylase HDAC8.
  JOURNAL   ACS Chem Biol. 2017 Sep 15;12(9):2281-2286. doi: 
            10.1021/acschembio.7b00330. Epub 2017 Aug 30.
  PUBMED    28846375
REFERENCE   2  (bases 1 to 7575)
  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: 
            "10.1021/acschembio.7b00330"; journalName: "ACS Chem Biol"; date: 
            "2017-09-15- 15"; volume: "12"; issue: "9"; pages: "2281-2286"
COMMENT     Sequence Label: pET His6 MBP TEV HDAC8_374
FEATURES             Location/Qualifiers
     source          1..7575
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     119..138
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     222..241
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        222..240
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    241..265
                     /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)."
     RBS             280..302
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             321..338
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             348..1448
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1422..1445
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1461..1481
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2660..2677
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2730..2748)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2744..2791
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2828..3283
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2915..2934)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3125..3146
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3376..4191)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(3376..4191)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4099..4118
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4183..4202
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4313..4901
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4802..4821
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5055..5072
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5087..5226
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5152..5171)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5312..5334
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5328..5519)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6328..7410)
                     /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     7372..7391
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7411..7488)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca
       61 tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg
      121 tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga
      181 tgcgtccggc gtagaggatc gagatctcga tcccgcgaaa ttaatacgac tcactatagg
      241 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga
      301 gatataccat gggttcttct caccatcacc atcaccatgg ttcttctatg aaaatcgaag
      361 aaggtaaact ggtaatctgg attaacggcg ataaaggcta taacggtctc gctgaagtcg
      421 gtaagaaatt cgagaaagat accggaatta aagtcaccgt tgagcatccg gataaactgg
      481 aagagaaatt cccacaggtt gcggcaactg gcgatggccc tgacattatc ttctgggcac
      541 acgaccgctt tggtggctac gctcaatctg gcctgttggc tgaaatcacc ccggacaaag
      601 cgttccagga caagctgtat ccgtttacct gggatgccgt acgttacaac ggcaagctga
      661 ttgcttaccc gatcgctgtt gaagcgttat cgctgattta taacaaagat ctgctgccga
      721 acccgccaaa aacctgggaa gagatcccgg cgctggataa agaactgaaa gcgaaaggta
      781 agagcgcgct gatgttcaac ctgcaagaac cgtacttcac ctggccgctg attgctgctg
      841 acgggggtta tgcgttcaag tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg
      901 ataacgctgg cgcgaaagcg ggtctgacct tcctggttga cctgattaaa aacaaacaca
      961 tgaatgcaga caccgattac tccatcgcag aagctgcctt taataaaggc gaaacagcga
     1021 tgaccatcaa cggcccgtgg gcatggtcca acatcgacac cagcaaagtg aattatggtg
     1081 taacggtact gccgaccttc aagggtcaac catccaaacc gttcgttggc gtgctgagcg
     1141 caggtattaa cgccgccagt ccgaacaaag agctggcaaa agagttcctc gaaaactatc
     1201 tgctgactga tgaaggtctg gaagcggtta ataaagacaa accgctgggt gccgtagcgc
     1261 tgaagtctta cgaggaagag ttggcgaaag atccacgtat tgccgccact atggaaaacg
     1321 cccagaaagg tgaaatcatg ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc
     1381 gtactgcggt gatcaacgcc gccagcggtc gtcagactgt cgatgaagcc ctgaaagacg
     1441 cgcagactaa tgggatcgag gaaaacctgt acttccaatc caatgcagac tctgggcagt
     1501 ctctggttcc ggtttacatc tattctccgg aatacgtttc tatgtgcgac tccctggcta
     1561 aaatcccgaa acgtgcatcg atggtgcatt ctttgattga agcatatgca ctgcataagc
     1621 agatgaggat agttaagcct aaagtggcct ccatggagga gatggccacc ttccacactg
     1681 atgcttatct gcagcatctc cagaaggtca gccaagaggg cgatgatgat catccggact
     1741 ccatagaata tgggctaggt tatgactgcc cagccactga agggatattt gactatgcag
     1801 cagctatagg aggggctacg atcacagctg cccaatgcct gattgacgga atgtgcaaag
     1861 tagcaattaa ctggtctgga gggtggcatc atgcaaagaa agatgaagct tctggtttct
     1921 gttacctgaa cgatgccgtt ctgggtatcc tgcgtctgcg tcgtaaattc gaacgtatcc
     1981 tgtacgttga tctggacctg caccacggtg atggtgttga agacgcgttc agtttcacct
     2041 ccaaagtcat gaccgtgtcc ctgcacaaat tctccccagg atttttccca ggaacaggtg
     2101 acgtgtctga tgttggccta gggaagggac ggtactacag tgtaaatgtg cccattcagg
     2161 atggcataca agatgaaaaa tattaccaga tctgtgaaag tgtactaaag gaagtatacc
     2221 aagcctttaa tcccaaagca gtggtcttac agctgggagc tgacacaata gctggggatc
     2281 ccatgtgctc ctttaacatg actccagtgg gaattggcaa gtgtcttaag tacatcctgc
     2341 agtggcagct ggctaccctg atcctgggtg gcggtggtta caacctggcg aacactgctc
     2401 gctgctggac ctacctgacc ggcgtaatcc tgggcaaaac cctgagctct gagatcccag
     2461 atcatgagtt tttcacagca tatggtcctg attatgtgct ggaaatcacg ccaagctgca
     2521 ggcctgaccg caacgagccg caccgcatcc agcagatcct caactacatc aaaggcaacc
     2581 tgaaatgagg taataacatt ggaagtggat aacggatccg aattcgagcg ccgtcgacaa
     2641 gcttgcggcc gcactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc
     2701 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     2761 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
     2821 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     2881 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     2941 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     3001 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     3061 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     3121 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     3181 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     3241 aaatttaacg cgaattttaa caaactagta acgtttacaa tttcaggtgg cacttttcgg
     3301 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     3361 ctcatgaatt aattcttaga aaaactcatc gagcatcaaa tgaaactgca atttattcat
     3421 atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc
     3481 accgaggcag ttccatagga tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc
     3541 aacatcaata caacctatta atttcccctc gtcaaaaata aggttatcaa gtgagaaatc
     3601 accatgagtg acgactgaat ccggtgagaa tggcaaaagt ttatgcattt ctttccagac
     3661 ttgttcaaca ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt
     3721 attcattcgt gattgcgcct gagcgagacg aaatacgcga tcgctgttaa aaggacaatt
     3781 acaaacagga atcgaatgca accggcgcag gaacactgcc agcgcatcaa caatgttttc
     3841 acctgaatca ggatattctt ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt
     3901 gagtaaccat gcatcatcag gagtacggat aaaatgcttg atggtcggaa gaggcataaa
     3961 ttccgtcagc cagtttagtc tgaccatctc atctgtaaca tcattggcaa cgctaccttt
     4021 gccatgtttc agaaacaact ctggcgcatc gggcttccca tacaatcgat agattgtcgc
     4081 acctgattgc ccgacattat cgcgagccca tttataccca tataaatcag catccatgtt
     4141 ggaatttaat cgcggcctag agcaagacgt ttcccgttga atatggctca taacacccct
     4201 tgtattactg tttatgtaag cagacagttt tattgttcat gaccaaaatc ccttaacgtg
     4261 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     4321 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     4381 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     4441 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     4501 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     4561 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     4621 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     4681 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     4741 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     4801 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     4861 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     4921 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     4981 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     5041 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt
     5101 ttctccttac gcatctgtgc ggtatttcac accgcaatgg tgcactctca gtacaatctg
     5161 ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg
     5221 gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     5281 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     5341 ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc
     5401 gattcacaga tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt
     5461 aatgtctggc ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact
     5521 gatgcctccg tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag
     5581 aggatgctca cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag
     5641 ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc
     5701 cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg
     5761 cagatccgga acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca
     5821 cggaaaccga agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg
     5881 cttcacgttc gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag
     5941 cctagccggg tcctcaacga caggagcacg atcatgcgca cccgtggggc cgccatgccg
     6001 gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga
     6061 gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt cgcgctccag
     6121 cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc tacgagttgc
     6181 atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc ccaccggaag
     6241 gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc ctaatgagtg
     6301 agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     6361 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     6421 cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg
     6481 gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg
     6541 tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac
     6601 taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag
     6661 cgccatctga tcgttggcaa ccagcatcgc agtgggaacg ataccctcat tcagcatttg
     6721 catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg
     6781 aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga
     6841 acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac
     6901 gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga
     6961 gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg
     7021 gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac
     7081 cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc
     7141 cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag
     7201 actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg
     7261 gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga
     7321 aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc
     7381 tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg
     7441 gcgctatcat gccataccgc gaaaggtttt gcgccattcg atggtgtccg ggatctcgac
     7501 gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga
     7561 gcaccgccgc cgcaa
//
