LOCUS       sequence-145743-        5986 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Sacsin Ubl-sr1 (SIRPT1 repeat) domain (1-339) in fusion 
            with GST (bacterial).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5986)
  AUTHORS   Menade M, Kozlov G, Trempe JF, Pande H, Shenker S, Wickremasinghe S,
            Li X, Hojjat H, Dicaire MJ, Brais B, McPherson PS, Wong MJH, Young 
            JC, Gehring K
  TITLE     Structures of ubiquitin-like (Ubl) and Hsp90-like domains of sacsin 
            provide insight into pathological mutations.
  JOURNAL   J Biol Chem. 2018 Aug 17;293(33):12832-12842. doi: 
            10.1074/jbc.RA118.003939. Epub 2018 Jun 26.
  PUBMED    29945973
REFERENCE   2  (bases 1 to 5986)
  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"; doi: 
            "10.1074/jbc.RA118.003939"; journalName: "J Biol Chem"; date: 
            "2018-08-17- 17"; volume: "293"; issue: "33"; pages: "12832-12842"
COMMENT     Sequence Label: pGEX6P1-Ubl-sr1(SIRPT1)
FEATURES             Location/Qualifiers
     source          1..5986
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..941
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(2036..2058)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2203..2221)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2289..2393
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2394..2462,2463..3254)
                     /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(2612..2631)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3425..4013
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3914..3933
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4167..4184
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4257..4334
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             4335..5417
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4354..4373)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    5430..5451
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5466..5471,5472..5489,5490..5496)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5504..5520
                     /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     5528..5544
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5528..5544
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5540..5986,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5560..5577)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5560..5576)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5569..5591)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gaagttctgt tccaggggcc cctgggatcc atggagacca
      961 aggagaacag gtgggtcccg gtgaccgtgc tccccggctg cgtgggctgc aggaccgtcg
     1021 cggcgctggc gtcctggacc gtgcgcgatg tgaaggaacg tatcttcgcg gagactggct
     1081 tcccggtgtc ggagcagcgg ctgtggcgcg gcggccgcga gttatctgac tggatcaaga
     1141 ttggagatct gacttccaaa aattgtcatc tttttgtaaa ccttcaatca aaaggcttaa
     1201 aagggggagg tcgatttggt cagacaacgc caccacttgt tgattttctc aaggacattt
     1261 tgagaagata tccagaagga ggacagattc ttaaggaatt aattcagaat gcagaagatg
     1321 ctggggcgac agaagttaaa tttttatatg atgaaactca atacggaaca gagactcttt
     1381 ggtcaaaaga tatggcgcca tatcaggggc cagctctcta tgtgtacaac aacgcggttt
     1441 tcaccccaga ggactggcac ggcattcaag aaatagcaag aagcaggaaa aaggatgatc
     1501 ctctgaaggt cggaagattt ggaattgggt ttaattctgt ctatcatata acagatgttc
     1561 cttgtatctt tagtggtgac caaatcggga tgctagatcc tcatcaaaca ctttttggcc
     1621 cacatgaatc aggccaatgt tggaatctca aagatgacag caaagaaatt agtgaacttt
     1681 cagaccagtt tgcaccattt gttggcattt ttggaagcac caaggaaaca tttataaacg
     1741 gcaattttcc aggaacattt ttccgtttcc ctcttcgcct acaaccttca caacttagta
     1801 gtaacctcta caataagcag aaggttcttg agttgtttga gtcttttagg gcagatgcgg
     1861 acacagtgct gctctttctg aaaagtgtgc aggatgtttc cttatatgtc cgagaggctg
     1921 acggaacaga gaaactggtg tttagagtga cttcgagtga gagtaagtag ctcgagcggc
     1981 cgcatcgtga ctgactgacg atctgcctcg cgcgtttcgg tgatgacggt gaaaacctct
     2041 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
     2101 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt
     2161 cacgtagcga tagcggagtg tataattctt gaagacgaaa gggcctcgtg atacgcctat
     2221 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
     2281 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2341 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     2401 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     2461 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     2521 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     2581 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtgttg
     2641 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     2701 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     2761 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     2821 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     2881 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag
     2941 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     3001 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     3061 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     3121 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     3181 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     3241 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     3301 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     3361 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     3421 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     3481 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     3541 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     3601 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     3661 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     3721 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     3781 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     3841 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     3901 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3961 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4021 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     4081 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     4141 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     4201 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata aattccgaca
     4261 ccatcgaatg gtgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat
     4321 tcagggtggt gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct
     4381 cttatcagac cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg
     4441 aaaaagtgga agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac
     4501 tggcgggcaa acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc
     4561 cgtcgcaaat tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg
     4621 tgtcgatggt agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg
     4681 cgcaacgcgt cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg
     4741 tggaagctgc ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca
     4801 tcaacagtat tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg
     4861 cattgggtca ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc
     4921 tgcgtctggc tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac
     4981 gggaaggcga ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg
     5041 gcatcgttcc cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg
     5101 ccattaccga gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata
     5161 ccgaagacag ctcatgttat atcccgccgt caaccaccat caaacaggat tttcgcctgc
     5221 tggggcaaac cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca
     5281 atcagctgtt gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa
     5341 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     5401 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     5461 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     5521 tttcacacag gaaacagcta tgaccatgat tacggattca ctggccgtcg ttttacaacg
     5581 tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt
     5641 cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
     5701 cctgaatggc gaatggcgct ttgcctggtt tccggcacca gaagcggtgc cggaaagctg
     5761 gctggagtgc gatcttcctg aggccgatac tgtcgtcgtc ccctcaaact ggcagatgca
     5821 cggttacgat gcgcccatct acaccaacgt aacctatccc attacggtca atccgccgtt
     5881 tgttcccacg gagaatccga cgggttgtta ctcgctcaca tttaatgttg atgaaagctg
     5941 gctacaggaa ggccagacgc gaattatttt tgatggcgtt ggaatt
//
