LOCUS       sequence-447917-        5109 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human KDAC8 (HDAC8) H143A in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5109)
  AUTHORS   Toro TB, Painter RG, Haynes RA, Glotser EY, Bratton MR, Bryant JR, 
            Nichols KA, Matthew-Onabanjo AN, Matthew AN, Bratcher DR, Perry CD, 
            Watt TJ
  TITLE     Purification of metal-dependent lysine deacetylases with 
            consistently high activity.
  JOURNAL   Protein Expr Purif. 2018 Jan;141:1-6. doi: 
            10.1016/j.pep.2017.08.009. Epub 2017 Aug 24.
  PUBMED    28843507
REFERENCE   2  (bases 1 to 5109)
  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: "Protein 
            Expr Purif."; date: "2018-01"; volume: "141:1-6. doi"; pages: " 
            10.1016/j.pep.2017.08.009. Epub 2017 Aug 24"
COMMENT     Sequence Label: pJExpress401-KDAC8 H143A
FEATURES             Location/Qualifiers
     source          1..5109
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      86..674
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     575..594
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     828..845
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     943..960
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     944..960
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1417..1433
                     /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        join(1458..1472,1473..1478,1479..1495,1496..1502)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    1478..1494
                     /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)."
     protein_bind    1508..1527
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     CDS             2714..2734
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2735..2752
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2957..2976)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     promoter        3043..3134
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3135..3944
                     /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             4027..5109
                     /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     complement(4046..4065)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ctcatgacca aaatccctta acgtgagtta cgcgcgcgtc gttccactga gcgtcagacc
       61 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
      121 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
      181 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
      241 tgtagccgta gttagcccac cacttcaaga actctgtagc accgcctaca tacctcgctc
      301 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
      361 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
      421 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
      481 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
      541 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
      601 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
      661 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
      721 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
      781 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
      841 gcgaggaagc ggaaggcgag agtagggaac tgccaggcat caaactaagc agaaggcccc
      901 tgacggatgg cctttttgcg tttctacaaa ctctttctgt gttgtaaaac gacggccagt
      961 cttaagctcg ggccccctgg gcggttctga taacgagtaa tcgttaatcc gcaaataacg
     1021 taaaaacccg cttcggcggg tttttttatg gggggagttt agggaaagag catttgtcag
     1081 aatatttaag ggcgcctgtc actttgcttg atatatgaga attatttaac cttataaatg
     1141 agaaaaaagc aacgcacttt aaataagata cgttgctttt tcgattgatg aacacctata
     1201 attaaactat tcatctatta tttatgattt tttgtatata caatatttct agtttgttaa
     1261 agagaattaa gaaaataaat ctcgaaaata ataaagggaa aatcagtttt tgatatcaaa
     1321 attatacatg tcaacgataa tacaaaatat aatacaaact ataagatgtt atcagtattt
     1381 attatgcatt tagaataaat tttgtgtcgc ccttaattgt gagcggataa caattacgag
     1441 cttcatgcac agtgaaatca tgaaaaattt atttgctttg tgagcggata acaattataa
     1501 tatgtggaat tgtgagcgct cacaattcca caacggtttc cctctagaaa taattttgtt
     1561 taacttttag gaggtaaaac atatggaaga gcctgaagag cctgcagaca gcggccaatc
     1621 cctggtgccg gtttacattt acagcccgga atatgtgagc atgtgcgata gcttggcgaa
     1681 aatccctaag cgcgcgagca tggtccacag cctgatcgag gcgtatgcgt tgcacaaaca
     1741 aatgcgtatt gtgaaaccga aagtggcatc tatggaggaa atggctacct ttcacacgga
     1801 cgcgtacctg caacatctgc agaaagtgag ccaagaaggt gacgatgatc acccggacag
     1861 cattgagtat ggcctgggtt acgattgtcc ggccacggag ggtattttcg actacgcggc
     1921 agcaatcggt ggtgcgacga tcaccgcggc acagtgtctg atcgatggca tgtgcaaggt
     1981 tgccatcaat tggagcggcg gttggcatgc agcgaagaaa gacgaagcat ccggtttctg
     2041 ctacctgaac gatgcggttc tgggcatcct gcgtctgcgc cgtaagtttg aacgcattct
     2101 gtacgttgac ctggatctgc atcacggtga tggcgtcgag gacgccttct cctttacgag
     2161 caaagtcatg accgtgtctc tgcacaagtt cagcccgggt ttctttccgg gtaccggcga
     2221 tgttagcgat gttggtctgg gtaagggccg ttactacagc gtgaacgttc cgattcagga
     2281 tggtatccag gacgagaaat actatcagat ctgcgaaagc gtgctgaagg aagtttacca
     2341 ggctttcaat ccgaaagccg ttgtcttgca gctgggtgcc gacacgatcg caggtgaccc
     2401 gatgtgcagc ttcaatatga ccccggtggg tattggtaag tgtttgaaat acattctgca
     2461 atggcagctg gcgacgctga ttctgggtgg cggtggctac aacttggcta acaccgcgcg
     2521 ttgttggacc tatttgaccg gtgtcatcct gggcaagacc ctgagctcgg aaattccgga
     2581 ccacgagttt ttcaccgctt atggcccgga ttatgtactg gagatcactc cgtcttgccg
     2641 tccggaccgc aacgagccac accgtattca acaaattctg aactatatca aaggtaatct
     2701 gaagcacgtc gtcgagaatc tgtattttca gggccatcat catcaccatc actaactcga
     2761 gccccaaggg cgacaccccc taattagccc gggcgaaagg cccagtcttt cgactgagcc
     2821 tttcgtttta tttgatgcct ggcagttccc tactctcgca tggggagtcc ccacactacc
     2881 atcggcgcta cggcgtttca cttctgagtt cggcatgggg tcaggtggga ccaccgcgct
     2941 actgccgcca ggcaaacaag gggtgttatg agccatattc aggtataaat gggctcgcga
     3001 taatgttcag aattggttaa ttggttgtaa cactgacccc tatttgttta tttttctaaa
     3061 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     3121 gaaaaaggaa gaatatgagc catattcaac gggaaacgtc gaggccgcga ttaaattcca
     3181 acatggatgc tgatttatat gggtataaat gggctcgcga taatgtcggg caatcaggtg
     3241 cgacaatcta tcgcttgtat gggaagcccg atgcgccaga gttgtttctg aaacatggca
     3301 aaggtagcgt tgccaatgat gttacagatg agatggtcag actaaactgg ctgacggaat
     3361 ttatgccact tccgaccatc aagcatttta tccgtactcc tgatgatgca tggttactca
     3421 ccactgcgat ccccggaaaa acagcgttcc aggtattaga agaatatcct gattcaggtg
     3481 aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt gcactcgatt cctgtttgta
     3541 attgtccttt taacagcgat cgcgtatttc gcctcgctca ggcgcaatca cgaatgaata
     3601 acggtttggt tgatgcgagt gattttgatg acgagcgtaa tggctggcct gttgaacaag
     3661 tctggaaaga aatgcataaa cttttgccat tctcaccgga ttcagtcgtc actcatggtg
     3721 atttctcact tgataacctt atttttgacg aggggaaatt aataggttgt attgatgttg
     3781 gacgagtcgg aatcgcagac cgataccagg atcttgccat cctatggaac tgcctcggtg
     3841 agttttctcc ttcattacag aaacggcttt ttcaaaaata tggtattgat aatcctgata
     3901 tgaataaatt gcaatttcat ttgatgctcg atgagttttt ctaagcggcg cgccatcgaa
     3961 tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg
     4021 gtgaatatga aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag
     4081 accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg
     4141 gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc
     4201 aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa
     4261 attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg
     4321 gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc
     4381 gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct
     4441 gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt
     4501 attattttct cccatgagga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt
     4561 caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg
     4621 gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc
     4681 gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt
     4741 cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc
     4801 gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagat
     4861 agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa
     4921 accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg
     4981 ttgccagtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct
     5041 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     5101 gggcagtga
//
