LOCUS       sequence-445584-        6307 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of USP27X (Y381H) with GST tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6307)
  AUTHORS   Koch I, Slovik M, Zhang Y, Liu B, Rennie M, Konz E, Cogne B, Daana 
            M, Davids L, Diets IJ, Gold NB, Holtz AM, Isidor B, Mor-Shaked H, 
            Neira Fresneda J, Niederhoffer KY, Nizon M, Pfundt R, Simon M, 
            Stegmann A, Guillen Sacoto MJ, Wevers M, Barakat TS, Yanovsky-Dagan 
            S, Atanassov BS, Toth R, Gao C, Bustos F, Harel T
  TITLE     USP27X variants underlying X-linked intellectual disability disrupt 
            protein function via distinct mechanisms.
  JOURNAL   Life Sci Alliance. 2024 Jan 5;7(3):e202302258. doi: 
            10.26508/lsa.202302258. Print 2024 Mar.
  PUBMED    38182161
REFERENCE   2  (bases 1 to 6307)
  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: "Life Sci 
            Alliance."; date: "2024-01-5"; volume: "7(3):e202302258. doi"; 
            pages: " 10.26508/lsa.202302258. Print 2024 Mar"
COMMENT     Sequence Label: pGEX6P1 GST-hUSP27X Y381H
FEATURES             Location/Qualifiers
     source          1..6307
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(575..580,581..598,599..605)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    613..629
                     /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)."
     primer_bind     637..653
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     637..653
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             649..1176
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(669..686)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(669..685)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(678..700)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(1278..1283,1284..1299,1300..1306)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1314..1330
                     /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             1353..2006
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1964..1986
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2013..2036
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(3452..3474)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3619..3637)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3705..3809
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3810..3878,3879..4670)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4028..4047)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4841..5429
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5330..5349
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5583..5600
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        5673..5750
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(5751..6307,1..526)
                     /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(5770..5789)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gggcccatta agttctgtct cggcgcgtct gcgtctggct ggctggcata aatatctcac
       61 tcgcaatcaa attcagccga tagcggaacg ggaaggcgac tggagtgcca tgtccggttt
      121 tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc actgcgatgc tggttgccaa
      181 cgatcagatg gcgctgggcg caatgcgcgc cattaccgag tccgggctgc gcgttggtgc
      241 ggatatctcg gtagtgggat acgacgatac cgaagacagc tcatgttata tcccgccgtt
      301 aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca
      361 actctctcag ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag
      421 aaaaaccacg ctagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
      481 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
      541 atgtaagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
      601 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
      661 acggattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
      721 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
      781 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgctt tgcctggttt
      841 ccggcaccag aagcggtgcc ggaaagctgg ctggagtgcg atcttcctga ggccgatact
      901 gtcgtcgtcc cctcaaactg gcagatgcac ggttacgatg cgcccatcta caccaacgtg
      961 acctatccca ttacggtcaa tccgccgttt gttcccacgg agaatccgac gggttgttac
     1021 tcgctcacat ttaatgttga tgaaagctgg ctacaggaag gccagacgcg aattattttt
     1081 gatggcgttg gaattagctt atcgactgca cggtgcacca atgcttctgg cgtcaggcag
     1141 ccatcggaag ctgtggtatg gctgtgcagg tcgtaaatca ctgcataatt cgtgtcgctc
     1201 aaggcgcact cccgttctgg ataatgtttt ttgcgccgac atcataacgg ttctggcaaa
     1261 tattctgaaa tgagctgttg acaattaatc atcggctcgt ataatgtgtg gaattgtgag
     1321 cggataacaa tttcacacag gaaacagtat tcatgtcccc tatactaggt tattggaaaa
     1381 ttaagggcct tgtgcaaccc actcgacttc ttttggaata tcttgaagaa aaatatgaag
     1441 agcatttgta tgagcgcgat gaaggtgata aatggcgaaa caaaaagttt gaattgggtt
     1501 tggagtttcc caatcttcct tattatattg atggtgatgt taaattaaca cagtctatgg
     1561 ccatcatacg ttatatagct gacaagcaca acatgttggg tggttgtcca aaagagcgtg
     1621 cagagatttc aatgcttgaa ggagcggttt tggatattag atacggtgtt tcgagaattg
     1681 catatagtaa agactttgaa actctcaaag ttgattttct tagcaagcta cctgaaatgc
     1741 tgaaaatgtt cgaagatcgt ttatgtcata aaacatattt aaatggtgat catgtaaccc
     1801 atcctgactt catgttgtat gacgctcttg atgttgtttt atacatggac ccaatgtgcc
     1861 tggatgcgtt cccaaaatta gtttgtttta aaaaacgtat tgaagctatc ccacaaattg
     1921 ataagtactt gaaatccagc aagtatatag catggccttt gcagggctgg caagccacgt
     1981 ttggtggtgg cgaccatcct ccaaaatcgg atctggaagt tctgttccag gggcccctgg
     2041 gatccatgtg taaggactat gtatatgaca aagacattga gcaaattgcc aaagaagagc
     2101 aaggagaagc tttgaaatta caagcctcca cctcaacaga ggtttctcac cagcagtgtt
     2161 cagtgccagg ccttggtgag aaattcccaa cctgggaaac aaccaaacca gaattagaac
     2221 tgctggggca caacccgagg agaagaagaa tcacctccag ctttacgatc ggtttaagag
     2281 gactcatcaa tcttggcaac acgtgcttta tgaactgcat tgtccaggcc ctcacccaca
     2341 cgccgatact gagagatttc tttctctctg acaggcaccg atgtgagatg ccgagtcccg
     2401 agttgtgtct ggtctgtgag atgtcgtcgc tgtttcggga gttgtattct ggaaacccgt
     2461 ctcctcatgt gccctataag ttactgcacc tggtgtggat acatgcccgc catttagcag
     2521 ggtacaggca acaggatgcc cacgagttcc tcattgcagc gttagatgtc ctgcacaggc
     2581 actgcaaagg tgatgatgtc gggaaggcgg ccaacaatcc caaccactgt aactgcatca
     2641 tagaccaaat cttcacaggt ggcctgcagt ctgatgtcac ctgtcaagcc tgccatggcg
     2701 tctccaccac gatagaccca tgctgggaca ttagtttgga cttgcctggc tcttgcacct
     2761 ccttctggcc catgagccca gggagggaga gcagtgtgaa cggggaaagc cacataccag
     2821 gaatcaccac cctcacggac tgcttgcgga ggtttacgag gccagagcac ttaggaagca
     2881 gtgccaaaat caaatgtggt agttgccaaa gctaccagga atctaccaaa cagctcacaa
     2941 tgaataaatt acctgtcgtt gcctgttttc atttcaaacg gtttgaacat tcagcgaaac
     3001 agaggcgcaa gatcactaca tacatttcct ttcctctgga gctggatatg acgccgttta
     3061 tggcctcaag taaagagagc agaatgaatg gacaattgca gctgccaacc aatagtggaa
     3121 acaacgaaaa taagtattcc ttgtttgctg tggttaatca ccaaggaacc ttggagagtg
     3181 gccaccatac cagcttcatc cggcaccaca aggaccagtg gttcaagtgt gatgatgccg
     3241 tcatcactaa ggccagtatt aaggacgtac tggacagtga agggtattta ctgttctatc
     3301 acaaacaggt gctagaacat gagtcagaaa aagtgaaaga aatgaacaca caagcctact
     3361 gaggatcccc ggaattcccg ggtcgactcg agcggccgca tcgtgactga ctgacgatct
     3421 gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     3481 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
     3541 gtgttggcgg gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata
     3601 attcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     3661 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     3721 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     3781 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     3841 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     3901 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     3961 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     4021 aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag caactcggtc
     4081 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     4141 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     4201 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     4261 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     4321 taccaaacga cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac
     4381 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     4441 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     4501 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     4561 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     4621 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     4681 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     4741 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     4801 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     4861 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4921 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     4981 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     5041 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     5101 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     5161 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     5221 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     5281 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     5341 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     5401 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     5461 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     5521 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     5581 gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc
     5641 atctgtgcgg tatttcacac cgcataaatt ccgacaccat cgaatggcgc aaaacctttc
     5701 gcggtatggc atgatagcgc ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag
     5761 taacgttata cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg
     5821 tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg
     5881 agctgaatta cattcccaac cgcgtggcac aacaactggc gggcaaacag tcgttgctga
     5941 ttggcgttgc cacctccagt ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta
     6001 aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg
     6061 tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca
     6121 ttaactatcc gctggatgac caggatgcca ttgctgtgga agctgcctgc actaatgttc
     6181 cggcgttatt tcttgatgtc tctgaccaga cacccatcaa cagtattatt ttctcccatg
     6241 aagacggtac gcgactgggc gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc
     6301 tgttagc
//
