LOCUS       sequence-445600-        7909 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transfection of USP27X (Y381H) with HA tag in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7909)
  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 7909)
  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: pCAGGS puro HA-hUSP27X Y381H
FEATURES             Location/Qualifiers
     source          1..7909
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1021..1040
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1021..1039
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     enhancer        1049..1428
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          1756..2772
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2696..2718
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2780..2799
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             2837..2863
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    4546..4680
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4583..4602)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4637..4656
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        4685..5184
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(4720..4743)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     5117..5136
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             5200..5799
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5200..5219)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5696..5716
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(6135..6152)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6135..6151)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6144..6166)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(6292..6747)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6429..6450)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6641..6660
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6774..6878
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6879..6947,6948..7739)
                     /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     complement(7097..7116)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga catgattacg aatttggcca agtcggcctc
     1021 taatacgact cactataggg agctcgtcga cattgattat tgactagtta ttaatagtaa
     1081 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg
     1141 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
     1201 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
     1261 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
     1321 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
     1381 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc
     1441 cccacgttct gcttcactct ccccatctcc cccccctccc cacccccaat tttgtattta
     1501 tttatttttt aattattttg tgcagcgatg ggggcggggg gggggggggg gtatgtgtag
     1561 gtgggggggg gggggggggg gggggggggg gggggggggg gcgggcgggg gggggcgggg
     1621 cggggcgagg ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa tcagagcggc
     1681 gcgctccgaa agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga
     1741 agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg ctccgccgcc
     1801 gcctcgcgcc gcccgccccg gctctgactg accgcgttac tcccacaggt gagcgggcgg
     1861 gacggccctt ctcctccggg ctgtaattag cgcttggttt aatgacggct tgtttctttt
     1921 ctgtggctgc gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg ggggagcggc
     1981 tcggggggtg cgtgcgtgtg tgtgtgcgtg gggagcgccg cgtgcggctc cgcgctgccc
     2041 ggcggctgtg agcgctgcgg gcgcggcgcg gggctttgtg cgctccgcag tgtgcgcgag
     2101 gggagcgcgg ccgggggcgg tgccccgcgg tgcggggggg gctgcgaggg gaacaaaggc
     2161 tgcgtgcggg gtgtgtgcgt gggggggtga gcagggggtg tgggcgcgtc ggtcgggctg
     2221 caaccccccc tgcacccccc tccccgagtt gctgagcacg gcccggcttc gggtgcgggg
     2281 ctccgtacgg ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg gcaggtgggg
     2341 gtgccgggcg gggcggggcc gcctcgggcc ggggagggct cgggggaggg gcgcggcggc
     2401 ccccggagcg ccggcggctg tcgaggcgcg gcgagccgca gccattgcct tttatggtaa
     2461 tcgtgcgaga gggcgcaggg acttcctttg tcccaaatct gtgcggagcc gaaatctggg
     2521 aggcgccgcc gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag
     2581 gaaatgggcg gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag
     2641 cctcggggct gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc
     2701 ggcttctggc gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc
     2761 tttttcctac agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa
     2821 gaattcaact cgaatgtacc catacgatgt gccagattac gccggatcca tgtgtaagga
     2881 ctatgtatat gacaaagaca ttgagcaaat tgccaaagaa gagcaaggag aagctttgaa
     2941 attacaagcc tccacctcaa cagaggtttc tcaccagcag tgttcagtgc caggccttgg
     3001 tgagaaattc ccaacctggg aaacaaccaa accagaatta gaactgctgg ggcacaaccc
     3061 gaggagaaga agaatcacct ccagctttac gatcggttta agaggactca tcaatcttgg
     3121 caacacgtgc tttatgaact gcattgtcca ggccctcacc cacacgccga tactgagaga
     3181 tttctttctc tctgacaggc accgatgtga gatgccgagt cccgagttgt gtctggtctg
     3241 tgagatgtcg tcgctgtttc gggagttgta ttctggaaac ccgtctcctc atgtgcccta
     3301 taagttactg cacctggtgt ggatacatgc ccgccattta gcagggtaca ggcaacagga
     3361 tgcccacgag ttcctcattg cagcgttaga tgtcctgcac aggcactgca aaggtgatga
     3421 tgtcgggaag gcggccaaca atcccaacca ctgtaactgc atcatagacc aaatcttcac
     3481 aggtggcctg cagtctgatg tcacctgtca agcctgccat ggcgtctcca ccacgataga
     3541 cccatgctgg gacattagtt tggacttgcc tggctcttgc acctccttct ggcccatgag
     3601 cccagggagg gagagcagtg tgaacgggga aagccacata ccaggaatca ccaccctcac
     3661 ggactgcttg cggaggttta cgaggccaga gcacttagga agcagtgcca aaatcaaatg
     3721 tggtagttgc caaagctacc aggaatctac caaacagctc acaatgaata aattacctgt
     3781 cgttgcctgt tttcatttca aacggtttga acattcagcg aaacagaggc gcaagatcac
     3841 tacatacatt tcctttcctc tggagctgga tatgacgccg tttatggcct caagtaaaga
     3901 gagcagaatg aatggacaat tgcagctgcc aaccaatagt ggaaacaacg aaaataagta
     3961 ttccttgttt gctgtggtta atcaccaagg aaccttggag agtggccacc ataccagctt
     4021 catccggcac cacaaggacc agtggttcaa gtgtgatgat gccgtcatca ctaaggccag
     4081 tattaaggac gtactggaca gtgaagggta tttactgttc tatcacaaac aggtgctaga
     4141 acatgagtca gaaaaagtga aagaaatgaa cacacaagcc tactgatcga gccttaatta
     4201 agggctagca aatgcatgcg gccgccaatt ccagctgagc gccggtcgct accattacca
     4261 gttggtctgg tgtcaaaaat aataataacc gggcaggcca tgtctgcccg tatttcgcgt
     4321 aaggaaatcc attatgtact atttaaaaaa cacaaacttt tggatgttcg gtttattctt
     4381 tttcttttac ttttttatca tgggagccta cttcccgttt ttcccgattt ggctacatga
     4441 catcaaccat atcagcaaaa gtgatacggg tattattttt gccgctattt ctctgttctc
     4501 gctattattc caaccgctgt ttggtctgct ttctgacaaa ctcggaactt gtttattgca
     4561 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     4621 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc
     4681 taccgggtag gggaggcgct tttcccaagg cagtctggag catgcgcttt agcagccccg
     4741 ctgggcactt ggcgctacac aagtggcctc tggcctcgca cacattccac atccaccggt
     4801 aggcgccaac cggctccgtt ctttggtggc cccttcgcgc caccttctac tcctccccta
     4861 gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca ggacgtgaca aatggaagta
     4921 gcacgtctca ctagtctcgt gcagatggac agcaccgctg agcaatggaa gcgggtaggc
     4981 ctttggggca gcggccaata gcagctttgc tccttcgctt tctgggctca gaggctggga
     5041 aggggtgggt ccgggggcgg gctcaggggc gggctcaggg gcggggcggg cgcccgaagg
     5101 tcctccggag gcccggcatt ctgcacgctt caaaagcgca cgtctgccgc gctgttctcc
     5161 tcttcctcat ctccgggcct ttcgacaagc tagcttacca tgaccgagta caagcccacg
     5221 gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc
     5281 gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc
     5341 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     5401 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     5461 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     5521 cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     5581 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     5641 gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac
     5701 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     5761 ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac gcccgcccca cgacccgcag
     5821 cgcccgaccg aaaggagcgc acgaccccat gcatcgattt gccagccatc tgttgtttgc
     5881 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     5941 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     6001 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     6061 ggctctatgg cttctgaggc ggaaagaacc agctggggct cgacgcggcc ggccgctgac
     6121 caagtcagct tggcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     6181 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     6241 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg gaaattgtaa
     6301 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     6361 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     6421 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     6481 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     6541 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     6601 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     6661 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     6721 cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa
     6781 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     6841 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     6901 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     6961 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     7021 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     7081 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     7141 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     7201 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     7261 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     7321 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     7381 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     7441 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     7501 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     7561 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     7621 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     7681 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     7741 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     7801 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     7861 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttc
//
