LOCUS       sequence-206039-        5931 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for bacterial expression of His-tagged GST encoded by 
            non-preferred codons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5931)
  AUTHORS   Josse L, Sampson CDD, Tuite MF, Howland K, von der Haar T
  TITLE     Codon-Dependent Translational Accuracy Controls Protein Quality in 
            Escherichia coli but not in Saccharomyces cerevisiae
  JOURNAL   bioRxiv 200006
REFERENCE   2  (bases 1 to 5931)
  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: "bioRxiv 
            200006"
COMMENT     Sequence Label: pTH848-gst-disec
FEATURES             Location/Qualifiers
     source          1..5931
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     97..116
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        97..115
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    116..140
                     /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)."
     RBS             155..177
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             190..213
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             214..867
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="SPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKKF
                     ELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRYG
                     VSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVLY
                     MDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             892..909
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(962..980)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      976..1023
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1060..1515
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1147..1166)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1357..1378
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1607..2422)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1607..2422)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2330..2349
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2414..2433
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2544..3132
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3033..3052
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3286..3303
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3318..3457
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3383..3402)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3543..3565
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3559..3750)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4559..5641)
                     /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     5603..5622
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5642..5719)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     join(5925..5931,1..13)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt
       61 ccggcgtaga ggatcgagat ctcgatcccg cgaaattaat acgactcact ataggggaat
      121 tgtgagcgga taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat
      181 accatgggcc accatcacca tcaccatcac cattcaccca tactaggata ctggaagata
      241 aagggactag tacaacccac aaggctacta ctagagtacc tagaggagaa gtacgaggag
      301 cacctatacg agagggacga gggagacaag tggaggaata agaagttcga gctaggacta
      361 gagttcccca atctacccta ctacatagac ggagacgtaa agctaacaca atcaatggct
      421 ataataaggt acatagctga caagcacaat atgctaggag gatgtcccaa ggagagggct
      481 gagatatcaa tgctagaggg agctgtacta gacataaggt acggagtatc aaggatagct
      541 tactcaaagg acttcgagac actaaaggta gacttcctat caaagctacc cgagatgcta
      601 aagatgttcg aggacaggct atgtcacaag acatacctaa atggagacca cgtaacacac
      661 cccgacttca tgctatacga cgctctagac gtagtactat acatggaccc catgtgtcta
      721 gacgctttcc ccaagctagt atgtttcaag aagaggatag aggctatacc ccaaatagac
      781 aagtacctaa agtcatcaaa gtacatagct tggcccctac aaggatggca agctacattc
      841 ggaggaggag accacccccc caagtaataa aagcttgcgg ccgcactcga gcaccaccac
      901 caccaccact gagatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc
      961 accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt
     1021 ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt agcggcgcat
     1081 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     1141 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     1201 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     1261 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
     1321 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
     1381 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
     1441 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat
     1501 taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     1561 atttttctaa atacattcaa atatgtatcc gctcatgaat taattcttag aaaaactcat
     1621 cgagcatcaa atgaaactgc aatttattca tatcaggatt atcaatacca tatttttgaa
     1681 aaagccgttt ctgtaatgaa ggagaaaact caccgaggca gttccatagg atggcaagat
     1741 cctggtatcg gtctgcgatt ccgactcgtc caacatcaat acaacctatt aatttcccct
     1801 cgtcaaaaat aaggttatca agtgagaaat caccatgagt gacgactgaa tccggtgaga
     1861 atggcaaaag tttatgcatt tctttccaga cttgttcaac aggccagcca ttacgctcgt
     1921 catcaaaatc actcgcatca accaaaccgt tattcattcg tgattgcgcc tgagcgagac
     1981 gaaatacgcg atcgctgtta aaaggacaat tacaaacagg aatcgaatgc aaccggcgca
     2041 ggaacactgc cagcgcatca acaatatttt cacctgaatc aggatattct tctaatacct
     2101 ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca tgcatcatca ggagtacgga
     2161 taaaatgctt gatggtcgga agaggcataa attccgtcag ccagtttagt ctgaccatct
     2221 catctgtaac atcattggca acgctacctt tgccatgttt cagaaacaac tctggcgcat
     2281 cgggcttccc atacaatcga tagattgtcg cacctgattg cccgacatta tcgcgagccc
     2341 atttataccc atataaatca gcatccatgt tggaatttaa tcgcggccta gagcaagacg
     2401 tttcccgttg aatatggctc ataacacccc ttgtattact gtttatgtaa gcagacagtt
     2461 ttattgttca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     2521 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     2581 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     2641 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     2701 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     2761 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     2821 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     2881 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     2941 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     3001 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     3061 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     3121 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     3181 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     3241 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     3301 gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     3361 caccgcaatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtata
     3421 cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc caacacccgc
     3481 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     3541 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgaggcagct
     3601 gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct gttcatccgc
     3661 gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa agcgggccat
     3721 gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg gatttctgtt
     3781 catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg ttactgatga
     3841 tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat ggatgcggcg
     3901 ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag atgtaggtgt
     3961 tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg tgcagggcgc
     4021 tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc atgttgttgc
     4081 tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta tcggtgattc
     4141 attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg acaggagcac
     4201 gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg
     4261 tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc
     4321 aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca
     4381 gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc
     4441 gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg
     4501 catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc
     4561 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     4621 cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga
     4681 cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca
     4741 cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac
     4801 atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc
     4861 cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg
     4921 cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac
     4981 tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc
     5041 agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt
     5101 gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga
     5161 aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag
     5221 tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc
     5281 cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc
     5341 gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg
     5401 cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg
     5461 actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg
     5521 ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg
     5581 aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca
     5641 cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt
     5701 tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg
     5761 aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc
     5821 aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa
     5881 caagcgctca tgagcccgaa gtggcgagcc cgatcttccc catcggtgat g
//
