LOCUS       sequence-450060-        6278 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of USP27X (K181E) with GST tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6278)
  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 6278)
  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 K181E
FEATURES             Location/Qualifiers
     source          1..6278
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1378..1400)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1545..1563)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1631..1735
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1736..1804,1805..2596)
                     /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(1954..1973)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2767..3355
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3256..3275
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3509..3526
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3599..3676
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3677..4759
                     /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(3696..3715)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4808..4813,4814..4831,4832..4838)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4846..4862
                     /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     4870..4886
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4870..4886
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             4882..5409
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4902..4919)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4902..4918)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4911..4933)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(5511..5516,5517..5532,5533..5539)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    5547..5563
                     /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             5586..6239
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     6197..6219
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             6246..6269
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
ORIGIN
        1 atgtgtaagg actatgtata tgacaaagac attgagcaaa ttgccaaaga agagcaagga
       61 gaagctttga aattacaagc ctccacctca acagaggttt ctcaccagca gtgttcagtg
      121 ccaggccttg gtgagaaatt cccaacctgg gaaacaacca aaccagaatt agaactgctg
      181 gggcacaacc cgaggagaag aagaatcacc tccagcttta cgatcggttt aagaggactc
      241 atcaatcttg gcaacacgtg ctttatgaac tgcattgtcc aggccctcac ccacacgccg
      301 atactgagag atttctttct ctctgacagg caccgatgtg agatgccgag tcccgagttg
      361 tgtctggtct gtgagatgtc gtcgctgttt cgggagttgt attctggaaa cccgtctcct
      421 catgtgccct ataagttact gcacctggtg tggatacatg cccgccattt agcagggtac
      481 aggcaacagg atgcccacga gttcctcatt gcagcgttag atgtcctgca caggcactgc
      541 gaaggtgatg atgtcgggaa ggcggccaac aatcccaacc actgtaactg catcatagac
      601 caaatcttca caggtggcct gcagtctgat gtcacctgtc aagcctgcca tggcgtctcc
      661 accacgatag acccatgctg ggacattagt ttggacttgc ctggctcttg cacctccttc
      721 tggcccatga gcccagggag ggagagcagt gtgaacgggg aaagccacat accaggaatc
      781 accaccctca cggactgctt gcggaggttt acgaggccag agcacttagg aagcagtgcc
      841 aaaatcaaat gtggtagttg ccaaagctac caggaatcta ccaaacagct cacaatgaat
      901 aaattacctg tcgttgcctg ttttcatttc aaacggtttg aacattcagc gaaacagagg
      961 cgcaagatca ctacatacat ttcctttcct ctggagctgg atatgacgcc gtttatggcc
     1021 tcaagtaaag agagcagaat gaatggacaa ttgcagctgc caaccaatag tggaaacaac
     1081 gaaaataagt attccttgtt tgctgtggtt aatcaccaag gaaccttgga gagtggccac
     1141 tataccagct tcatccggca ccacaaggac cagtggttca agtgtgatga tgccgtcatc
     1201 actaaggcca gtattaagga cgtactggac agtgaagggt atttactgtt ctatcacaaa
     1261 caggtgctag aacatgagtc agaaaaagtg aaagaaatga acacacaagc ctactgagcg
     1321 gccgcatcgt gactgactga cgatctgcct cgcgcgtttc ggtgatgacg gtgaaaacct
     1381 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag
     1441 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca
     1501 gtcacgtagc gatagcggag tgtataattc ttgaagacga aagggcctcg tgatacgcct
     1561 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     1621 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     1681 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     1741 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     1801 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     1861 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     1921 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt
     1981 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     2041 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     2101 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     2161 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     2221 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgc
     2281 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     2341 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     2401 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     2461 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     2521 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     2581 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     2641 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     2701 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     2761 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2821 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     2881 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
     2941 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     3001 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     3061 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     3121 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3181 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3241 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3301 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3361 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     3421 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     3481 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     3541 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca taaattccga
     3601 caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca
     3661 attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt
     3721 ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg
     3781 ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca
     3841 actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc
     3901 gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt
     3961 ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct
     4021 cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc
     4081 tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc
     4141 catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt
     4201 cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg
     4261 tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga
     4321 acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga
     4381 gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg
     4441 cgccattacc gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga
     4501 taccgaagac agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct
     4561 gctggggcaa accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg
     4621 caatcagctg ttgcccgtct cactggtgaa aagaaaaacc acgctagcgc ccaatacgca
     4681 aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg
     4741 actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac
     4801 cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac
     4861 aatttcacac aggaaacagc tatgaccatg attacggatt cactggccgt cgttttacaa
     4921 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     4981 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     5041 agcctgaatg gcgaatggcg ctttgcctgg tttccggcac cagaagcggt gccggaaagc
     5101 tggctggagt gcgatcttcc tgaggccgat actgtcgtcg tcccctcaaa ctggcagatg
     5161 cacggttacg atgcgcccat ctacaccaac gtgacctatc ccattacggt caatccgccg
     5221 tttgttccca cggagaatcc gacgggttgt tactcgctca catttaatgt tgatgaaagc
     5281 tggctacagg aaggccagac gcgaattatt tttgatggcg ttggaattag cttatcgact
     5341 gcacggtgca ccaatgcttc tggcgtcagg cagccatcgg aagctgtggt atggctgtgc
     5401 aggtcgtaaa tcactgcata attcgtgtcg ctcaaggcgc actcccgttc tggataatgt
     5461 tttttgcgcc gacatcataa cggttctggc aaatattctg aaatgagctg ttgacaatta
     5521 atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca caggaaacag
     5581 tattcatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa cccactcgac
     5641 ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc gatgaaggtg
     5701 ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt ccttattata
     5761 ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata gctgacaagc
     5821 acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt gaaggagcgg
     5881 ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt gaaactctca
     5941 aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat cgtttatgtc
     6001 ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg tatgacgctc
     6061 ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa ttagtttgtt
     6121 ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc agcaagtata
     6181 tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat cctccaaaat
     6241 cggatctgga agttctgttc caggggcccc tgggatcc
//
