LOCUS       sequence-212283-        5856 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression for structure determination; may not be full 
            ORF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5856)
  TITLE     SGC Oxford April 2018 plasmids
REFERENCE   2  (bases 1 to 5856)
  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"
COMMENT     Sequence Label: KEAP1 (KEAP1A-c029)
FEATURES             Location/Qualifiers
     source          1..5856
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             92..109
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             134..154
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             721..738
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(791..809)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      805..852
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      889..1344
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(976..995)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1186..1207
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1436..2251)
                     /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(1436..2251)
                     /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     2159..2178
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2243..2262
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2373..2961
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2862..2881
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3115..3132
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3147..3286
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3212..3231)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3372..3394
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3388..3579)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4388..5470)
                     /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     5432..5451
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5471..5548)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5754..5773
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gcaccatcat catcatcatt cttctggtgt
      121 agatctgggt accgagaacc tgtacttcca atccatgggc aaccgcacct tcagctacac
      181 cctggaggat cataccaagc aggcctttgg catcatgaac gagctgcggc tcagccagca
      241 gctgtgtgac gtcacactgc aggtcaagta ccaggatgca ccggccgccc agttcatggc
      301 ccacaaggtg gtgctggcct catccagccc tgtcttcaag gccatgttca ccaacgggct
      361 gcgggagcag ggcatggagg tggtgtccat tgagggtatc caccccaagg tcatggagcg
      421 cctcattgaa ttcgcctaca cggcctccat ctccatgggc gagaagtgtg tcctccacgt
      481 catgaacggt gctgtcatgt accagatcga cagcgttgtc cgtgcctgca gtgacttcct
      541 ggtgcagcag ctggacccca gcaatgccat cggcatcgcc aacttcgctg agcagattgg
      601 ctgtgtggag ttgcaccagc gtgcccggga gtacatctac atgcattttg gggagtgaca
      661 gtaaaggtgg atacggatcc gaattcgagc tccgtcgaca agcttgcggc cgcactcgag
      721 caccaccacc accaccactg agatccggct gctaacaaag cccgaaagga agctgagttg
      781 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
      841 aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta
      901 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
      961 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     1021 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     1081 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     1141 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     1201 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     1261 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     1321 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     1381 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgaatt aattcttaga
     1441 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
     1501 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
     1561 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
     1621 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
     1681 ccggtgagaa tggcaaaagt ttatgcattt ctttccagac ttgttcaaca ggccagccat
     1741 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
     1801 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
     1861 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
     1921 ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag
     1981 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
     2041 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
     2101 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
     2161 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctag
     2221 agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
     2281 cagacagttt tattgttcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     2341 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     2401 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     2461 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     2521 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     2581 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     2641 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     2701 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     2761 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     2821 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     2881 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     2941 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     3001 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     3061 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     3121 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc
     3181 ggtatttcac accgcaatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     3241 gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc
     3301 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     3361 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     3421 gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg
     3481 ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa
     3541 gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg
     3601 atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt
     3661 tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg
     3721 gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga
     3781 tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt
     3841 gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca
     3901 tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat
     3961 cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga
     4021 caggagcacg atcatgcgca cccgtggggc cgccatgccg gcgataatgg cctgcttctc
     4081 gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt gcaagattcc
     4141 gaataccgca agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa
     4201 aatgacccag agcgctgccg gcacctgtcc tacgagttgc atgataaaga agacagtcat
     4261 aagtgcggcg acgatagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc
     4321 tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc
     4381 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4441 cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca
     4501 ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca
     4561 agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg
     4621 ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa
     4681 cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa
     4741 ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg
     4801 acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat
     4861 atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca
     4921 gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt
     4981 catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg
     5041 gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa
     5101 tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga
     5161 cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt
     5221 taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa
     5281 tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct
     5341 ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca
     5401 ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta
     5461 ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc
     5521 gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc
     5581 tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat
     5641 ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc
     5701 acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg
     5761 tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt
     5821 ccggcgtaga ggatcgagat ctcgatcccg cgaaat
//
