LOCUS       sequence-494415-        5555 bp    DNA     circular SYN 02-DEC-2025
DEFINITION  Bacterial expression of human DNAJC5 L116 deleted with GST and a 
            cleavage site for TEV protease on its N-terminus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5555)
  AUTHORS   Lee J, Chin N, Zou J, Mazli WNAB, Jarnik M, Saidi L, Xu Y, Jeong E, 
            Suh J, Replogle J, Ward ME, Bonifacino JS, Zheng W, Hao L, Ye Y
  TITLE     CHIP protects lysosomes from CLN4 mutant-induced membrane damage.
  JOURNAL   Nat Cell Biol. 2025 Aug 25. doi: 10.1038/s41556-025-01738-2.
  PUBMED    40855364
REFERENCE   2  (bases 1 to 5555)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 2, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat Cell 
            Biol. 2025 Aug 25. doi: 10.1038/s41556-025-01738-2."
COMMENT     Sequence Label: pGEX GST-tev-hDNAJC5 delL116
FEATURES             Location/Qualifiers
     source          1..5555
                     /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             2019..2039
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     primer_bind     complement(2700..2722)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2867..2885)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2953..3057
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3058..3126,3127..3918)
                     /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(3276..3295)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4089..4677
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4578..4597
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4831..4848
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4921..4998
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(4999..5555,1..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(5018..5037)
                     /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 aaaaaccacc ctggcgccca 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 atactagtga aaacctgtac ttccaatccg
     2041 gttccgcaga ccagagacag cgctcactct ctacttccgg ggaatcgtta taccatgttc
     2101 ttggactgga caagaatgca acctcagatg acattaaaaa gtcctatcgg aagctggccc
     2161 tgaaatatca ccctgacaag aaccctgata acccagaggc tgcagacaag tttaaggaga
     2221 ttaacaacgc ccacgcaatc ctgacagacg ccacgaaaag aaacatttat gacaagtatg
     2281 gctcgctggg gctctatgtg gctgagcaat ttggggagga gaatgtcaac acctacttcg
     2341 tactctccag ctggtgggcc aaggcgctgt tcgtcttctg tggcctcacc tgctgctact
     2401 gctgctgctg tttatgctgt tgctttaact gctgctgtgg gaagtgcaag cccaaggcac
     2461 ctgagggtga agagacagaa ttctacgtat cccctgaaga cttggaggca cagctgcagt
     2521 ctgatgagag ggaggctaca gacacaccga tcgtcataca gccagcatcc gccacagaga
     2581 ccacccagct gacagctgac tcccacccca gctatcacac tgacgggttc aactaaaagc
     2641 ttaattcatc gtgactgact gacgatctgc ctcgcgcgtt tcggtgatga cggtgaaaac
     2701 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     2761 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc
     2821 cagtcacgta gcgatagcgg agtgtataat tcttgaagac gaaagggcct cgtgatacgc
     2881 ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt
     2941 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     3001 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     3061 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     3121 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     3181 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     3241 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     3301 gttgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     3361 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     3421 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     3481 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     3541 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     3601 gcagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     3661 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     3721 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
     3781 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     3841 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     3901 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
     3961 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     4021 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     4081 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     4141 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     4201 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     4261 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     4321 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     4381 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     4441 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     4501 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     4561 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     4621 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     4681 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     4741 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     4801 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag
     4861 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg cataaattcc
     4921 gacaccatcg aatggcgcaa aacctttcgc ggtatggcat gatagcgccc ggaagagagt
     4981 caattcaggg tggtgaatgt gaaaccagta acgttatacg atgtcgcaga gtatgccggt
     5041 gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg
     5101 cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg cgtggcacaa
     5161 caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct ggccctgcac
     5221 gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg
     5281 gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt gcacaatctt
     5341 ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca ggatgccatt
     5401 gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgtctc tgaccagaca
     5461 cccatcaaca gtattatttt ctcccatgaa gacggtacgc gactgggcgt ggagcatctg
     5521 gtcgcattgg gtcaccagca aatcgcgctg ttagc
//
