LOCUS       sequence-445596-        7886 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transfection of USP27X (G76S) with HA tag in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7886)
  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 7886)
  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 G76S
FEATURES             Location/Qualifiers
     source          1..7886
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     448..467
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     701..718
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    835..856
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(871..876,877..894,895..901)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    909..925
                     /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     914..936
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     933..949
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     933..949
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     979..998
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        979..997
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     enhancer        1007..1386
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1388..1663
                     /label=chicken beta-actin promoter
     intron          1664..2680
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2604..2626
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2688..2707
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             2744..2770
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    4481..4615
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4518..4537)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4572..4591
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        4620..5119
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(4655..4678)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     5052..5071
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             5135..5734
                     /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(5135..5154)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5631..5651
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(6070..6087)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6070..6086)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6079..6101)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(6227..6682)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6364..6385)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6576..6595
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6709..6813
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6814..6882,6883..7674)
                     /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(7032..7051)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(7845..7886,1..547)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
       61 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
      121 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
      181 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
      241 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
      301 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
      361 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
      421 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
      481 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
      541 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
      601 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
      661 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
      721 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
      781 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt
      841 gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt
      901 gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgaca tgattacgaa
      961 tttggccaag tcggcctcta atacgactca ctatagggag ctcgtcgaca ttgattattg
     1021 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
     1081 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     1141 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     1201 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
     1261 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
     1321 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
     1381 accatggtcg aggtgagccc cacgttctgc ttcactctcc ccatctcccc cccctcccca
     1441 cccccaattt tgtatttatt tattttttaa ttattttgtg cagcgatggg ggcggggggg
     1501 gggggggcgc gcgccaggcg gggcggggcg gggcgagggg cggggcgggg cgaggcggag
     1561 aggtgcggcg gcagccaatc agagcggcgc gctccgaaag tttcctttta tggcgaggcg
     1621 gcggcggcgg cggccctata aaaagcgaag cgcgcggcgg gcgggagtcg ctgcgcgctg
     1681 ccttcgcccc gtgccccgct ccgccgccgc ctcgcgccgc ccgccccggc tctgactgac
     1741 cgcgttactc ccacaggtga gcgggcggga cggcccttct cctccgggct gtaattagcg
     1801 cttggtttaa tgacggcttg tttcttttct gtggctgcgt gaaagccttg aggggctccg
     1861 ggagggccct ttgtgcgggg ggagcggctc ggggggtgcg tgcgtgtgtg tgtgcgtggg
     1921 gagcgccgcg tgcggctccg cgctgcccgg cggctgtgag cgctgcgggc gcggcgcggg
     1981 gctttgtgcg ctccgcagtg tgcgcgaggg gagcgcggcc gggggcggtg ccccgcggtg
     2041 cggggggggc tgcgagggga acaaaggctg cgtgcggggt gtgtgcgtgg gggggtgagc
     2101 agggggtgtg ggcgcgtcgg tcgggctgca accccccctg cacccccctc cccgagttgc
     2161 tgagcacggc ccggcttcgg gtgcggggct ccgtacgggg cgtggcgcgg ggctcgccgt
     2221 gccgggcggg gggtggcggc aggtgggggt gccgggcggg gcggggccgc ctcgggccgg
     2281 ggagggctcg ggggaggggc gcggcggccc ccggagcgcc ggcggctgtc gaggcgcggc
     2341 gagccgcagc cattgccttt tatggtaatc gtgcgagagg gcgcagggac ttcctttgtc
     2401 ccaaatctgt gcggagccga aatctgggag gcgccgccgc accccctcta gcgggcgcgg
     2461 ggcgaagcgg tgcggcgccg gcaggaagga aatgggcggg gagggccttc gtgcgtcgcc
     2521 gcgccgccgt ccccttctcc ctctccagcc tcggggctgt ccgcgggggg acggctgcct
     2581 tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg gctctagagc
     2641 ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca acgtgctggt
     2701 tattgtgctg tctcatcatt ttggcaaaga attcgccacc atgtacccat acgatgtgcc
     2761 agattacgcc ggatccatgt gtaaggacta tgtatatgac aaagacattg agcaaattgc
     2821 caaagaagag caaggagaag ctttgaaatt acaagcctcc acctcaacag aggtttctca
     2881 ccagcagtgt tcagtgccag gccttggtga gaaattccca acctgggaaa caaccaaacc
     2941 agaattagaa ctgctggggc acaacccgag gagaagaaga atcacctcca gctttacgat
     3001 cagtttaaga ggactcatca atcttggcaa cacgtgcttt atgaactgca ttgtccaggc
     3061 cctcacccac acgccgatac tgagagattt ctttctctct gacaggcacc gatgtgagat
     3121 gccgagtccc gagttgtgtc tggtctgtga gatgtcgtcg ctgtttcggg agttgtattc
     3181 tggaaacccg tctcctcatg tgccctataa gttactgcac ctggtgtgga tacatgcccg
     3241 ccatttagca gggtacaggc aacaggatgc ccacgagttc ctcattgcag cgttagatgt
     3301 cctgcacagg cactgcaaag gtgatgatgt cgggaaggcg gccaacaatc ccaaccactg
     3361 taactgcatc atagaccaaa tcttcacagg tggcctgcag tctgatgtca cctgtcaagc
     3421 ctgccatggc gtctccacca cgatagaccc atgctgggac attagtttgg acttgcctgg
     3481 ctcttgcacc tccttctggc ccatgagccc agggagggag agcagtgtga acggggaaag
     3541 ccacatacca ggaatcacca ccctcacgga ctgcttgcgg aggtttacga ggccagagca
     3601 cttaggaagc agtgccaaaa tcaaatgtgg tagttgccaa agctaccagg aatctaccaa
     3661 acagctcaca atgaataaat tacctgtcgt tgcctgtttt catttcaaac ggtttgaaca
     3721 ttcagcgaaa cagaggcgca agatcactac atacatttcc tttcctctgg agctggatat
     3781 gacgccgttt atggcctcaa gtaaagagag cagaatgaat ggacaattgc agctgccaac
     3841 caatagtgga aacaacgaaa ataagtattc cttgtttgct gtggttaatc accaaggaac
     3901 cttggagagt ggccactata ccagcttcat ccggcaccac aaggaccagt ggttcaagtg
     3961 tgatgatgcc gtcatcacta aggccagtat taaggacgta ctggacagtg aagggtattt
     4021 actgttctat cacaaacagg tgctagaaca tgagtcagaa aaagtgaaag aaatgaacac
     4081 acaagcctac tgaggatccg gcaagcttgg tacctcgcga ctcgagcctt aattaagggc
     4141 tagcaaatgc atgcggccgc caattccagc tgagcgccgg tcgctaccat taccagttgg
     4201 tctggtgtca aaaataataa taaccgggca ggccatgtct gcccgtattt cgcgtaagga
     4261 aatccattat gtactattta aaaaacacaa acttttggat gttcggttta ttctttttct
     4321 tttacttttt tatcatggga gcctacttcc cgtttttccc gatttggcta catgacatca
     4381 accatatcag caaaagtgat acgggtatta tttttgccgc tatttctctg ttctcgctat
     4441 tattccaacc gctgtttggt ctgctttctg acaaactcgg aacttgttta ttgcagctta
     4501 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     4561 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatctaccg
     4621 ggtaggggag gcgcttttcc caaggcagtc tggagcatgc gctttagcag ccccgctggg
     4681 cacttggcgc tacacaagtg gcctctggcc tcgcacacat tccacatcca ccggtaggcg
     4741 ccaaccggct ccgttctttg gtggcccctt cgcgccacct tctactcctc ccctagtcag
     4801 gaagttcccc cccgccccgc agctcgcgtc gtgcaggacg tgacaaatgg aagtagcacg
     4861 tctcactagt ctcgtgcaga tggacagcac cgctgagcaa tggaagcggg taggcctttg
     4921 gggcagcggc caatagcagc tttgctcctt cgctttctgg gctcagaggc tgggaagggg
     4981 tgggtccggg ggcgggctca ggggcgggct caggggcggg gcgggcgccc gaaggtcctc
     5041 cggaggcccg gcattctgca cgcttcaaaa gcgcacgtct gccgcgctgt tctcctcttc
     5101 ctcatctccg ggcctttcga caagctagct taccatgacc gagtacaagc ccacggtgcg
     5161 cctcgccacc cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga
     5221 ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct
     5281 gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga
     5341 cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc
     5401 cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat
     5461 ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg
     5521 cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga
     5581 ggcggccgag cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc
     5641 cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg
     5701 cacctggtgc atgacccgca agcccggtgc ctgacgcccg ccccacgacc cgcagcgccc
     5761 gaccgaaagg agcgcacgac cccatgcatc gatttgccag ccatctgttg tttgcccctc
     5821 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     5881 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     5941 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     6001 tatggcttct gaggcggaaa gaaccagctg gggctcgacg cggccggccg ctgaccaagt
     6061 cagcttggca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca
     6121 acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg
     6181 caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggaaat tgtaagcgtt
     6241 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     6301 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     6361 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     6421 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     6481 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     6541 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     6601 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     6661 aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     6721 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     6781 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     6841 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     6901 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     6961 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     7021 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     7081 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     7141 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     7201 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     7261 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     7321 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     7381 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     7441 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     7501 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     7561 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     7621 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     7681 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     7741 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     7801 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     7861 ctgcgcgtaa tctgctgctt gcaaac
//
