LOCUS       sequence-263349-        6127 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial overexpression and crystallization.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6127)
  AUTHORS   Nielson JR, Fredrickson EK, Waller TC, Rendon OZ, Schubert HL, Lin 
            Z, Hill CP, Rutter J
  TITLE     Sterol Oxidation Mediates Stress-Responsive Vms1 Translocation to 
            Mitochondria.
  JOURNAL   Mol Cell. 2017 Nov 16;68(4):673-685.e6. doi: 
            10.1016/j.molcel.2017.10.022.
  PUBMED    29149595
REFERENCE   2  (bases 1 to 6127)
  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: 
            "10.1016/j.molcel.2017.10"; journalName: "Mol Cell"; date: 
            "2017-11-16- 16"; volume: "68"; issue: "4"; pages: "673-685.e6"
COMMENT     Sequence Label: pGST-TEV-yVms1(1-417,delta(38-69))
FEATURES             Location/Qualifiers
     source          1..6127
                     /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             924..944
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(2177..2199)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2344..2362)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2430..2534
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2535..2603,2604..3395)
                     /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(2753..2772)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3566..4154
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4055..4074
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4308..4325
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4398..4475
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4476..5558
                     /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(4495..4514)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5607..5612,5613..5630,5631..5637)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5645..5661
                     /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     5669..5685
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5669..5685
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5681..6127,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5701..5718)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5701..5717)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5710..5732)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga 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 aatcgatacg accgaaaacc tgtattttca gggcgccatg gggatgaatt
      961 cacaaaaggc gagtaaaatg actggctcac taaaaaagaa tgatctttat atttttgatt
     1021 tgtctgagca acttctgaat tcattgaaat tgatgagctt tgacaatgtt atgcgttgta
     1081 gtgtttgtca aatgagcttt gattccagaa atgaacaaaa ggcccactac cagactgatt
     1141 atcatttgat gaatgttaag cgtaacctta gggggctgga tatcctatcc gttgaggaat
     1201 tcgatgcgct gatttctaag gagcatggta tcaagtctga agacgaaaat tctggcggcg
     1261 agcaaacttc aagcgatcac gaagaaagtg aggaagtcag cgatcgggac ccggacctac
     1321 aaactaataa ttatatggaa actataatcg aaaatgattt gcagaaatta gggttccaaa
     1381 aagacgagtc ggatgctatt agccatatta atacccaatc gccctacatt tatttcaaat
     1441 cgaagtatct tcaaaagaat gaggtcttgg cgatttataa gtctctattt aataaaagga
     1501 gtctttccaa ccccaatgaa gctttgactt tttggaattc gcaagagaac ccaatggcca
     1561 tttcagctct gtttatggtg ggaggtggac attttgcggg agcaattgtt tctcatcaaa
     1621 gactcaatgt caaggggaac gcgcacaaga aggacgaaac cttaattgag caagcggtca
     1681 atttcttaga gcacaagacg tttcacaggt atactaccag aaggaagcaa ggtggttctc
     1741 agtcagcaat ggacaatgca aagggaaaag ctaactctgc cggttcagca cttcgtaggt
     1801 acaatgaaag cgctttaaaa acagatatac agggtgtgtt gaaggactgg gaaccttatc
     1861 tgtcaaagtg tgataatatt tttattcgag ctcgcaatgt cagtgataaa aaaattttta
     1921 ctgataatac cgttctgaac aagggtgatg aaaggataaa gagcttccct tttaccacga
     1981 acaggcccac tgtattagaa ttgaagaagg catggtgcga actttcctac ctgaaaatat
     2041 tacctaagcc tgaaccattg gcagtgaaag aaacagtaca aaagttagaa gtgtcgaaca
     2101 aaaaggacta actcgagcgg ccgcatcgtg actgactgac gatctgcctc gcgcgtttcg
     2161 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     2221 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     2281 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtataattct tgaagacgaa
     2341 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     2401 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     2461 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     2521 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     2581 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     2641 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     2701 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     2761 gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt
     2821 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     2881 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     2941 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     3001 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     3061 gtgacaccac gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac
     3121 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     3181 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     3241 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     3301 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     3361 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     3421 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     3481 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     3541 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     3601 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     3661 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     3721 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     3781 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     3841 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     3901 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     3961 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     4021 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     4081 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     4141 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     4201 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     4261 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     4321 gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     4381 cacaccgcat aaattccgac accatcgaat ggcgcaaaac ctttcgcggt atggcatgat
     4441 agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg
     4501 tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc
     4561 acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc
     4621 ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct
     4681 ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc
     4741 aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag
     4801 cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg
     4861 atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg
     4921 atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac
     4981 tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat
     5041 taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc
     5101 aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa
     5161 ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga
     5221 tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct
     5281 cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca
     5341 tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc
     5401 agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca
     5461 ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     5521 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt
     5581 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt
     5641 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacggattc
     5701 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
     5761 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
     5821 cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc tttgcctggt ttccggcacc
     5881 agaagcggtg ccggaaagct ggctggagtg cgatcttcct gaggccgata ctgtcgtcgt
     5941 cccctcaaac tggcagatgc acggttacga tgcgcccatc tacaccaacg tgacctatcc
     6001 cattacggtc aatccgccgt ttgttcccac ggagaatccg acgggttgtt actcgctcac
     6061 atttaatgtt gatgaaagct ggctacagga aggccagacg cgaattattt ttgatggcgt
     6121 tggaatt
//
