LOCUS       sequence-319833-        5597 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5597)
  TITLE     SGC Toronto 2019-Q1 to 2020-Q2
REFERENCE   2  (bases 1 to 5597)
  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: 6WAJ
FEATURES             Location/Qualifiers
     source          1..5597
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(279..299)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(312..329)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             339..361
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    376..400
                     /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     complement(400..419)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(401..419)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(503..522)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        728..805
                     /gene="lacI"
                     /label=lacI promoter
     CDS             806..1888
                     /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(825..844)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             2697..2888
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2882..2904)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2990..3129
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3045..3064
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3144..3161)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3315..3903)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3395..3414)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4014..4033)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4025..4840
                     /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             4025..4840
                     /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     complement(4098..4117)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(4932..5387)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5069..5090)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5281..5300
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(5424..5471)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     5467..5485
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(5538..5555)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 tggtagatga tgtctaggac cttctctgtc tccactgcct gggactccaa cgtcttcccc
       61 agtgaggaga cgatctcagc atcgtggaca aagaaagcca gtggcagggg atcctcctgg
      121 gccagtagcg cgttgcacac gagctgcagc ctgtccgggg tgatgtccac gggcacgtcg
      181 aacggggaac ccagcagctg cccgccctca tcctggaact gcactagcaa ccgctgcaca
      241 tcgcgcgcca ccgcctcgtc cgccactgct gccgccatgc cctggaaata caagttttct
      301 ctgccgctgc tgtgatgatg atgatgatgc atggtatatc tccttcttaa agttaaacaa
      361 aattatttct agaggggaat tgttatccgc tcacaattcc cctatagtga gtcgtattaa
      421 tttcgcggga tcgagatctc gatcctctac gccggacgca tcgtggccgg catcaccggc
      481 gccacaggtg cggttgctgg cgcctatatc gccgacatca ccgatgggga agatcgggct
      541 cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg ccccgtggcc
      601 gggggactgt tgggcgccat ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac
      661 ggcctcaacc tactactggg ctgcttccta atgcaggagt cgcataaggg agagcgtcga
      721 gatcccggac accatcgaat ggcgcaaaac ctttcgcggt atggcatgat agcgcccgga
      781 agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg tcgcagagta
      841 tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc
      901 gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt
      961 ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc
     1021 cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc
     1081 cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca
     1141 caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga
     1201 tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga
     1261 ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac tgggcgtgga
     1321 gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt
     1381 ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc
     1441 gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat
     1501 gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg
     1561 cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg
     1621 atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga
     1681 ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc
     1741 ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc
     1801 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     1861 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt tagctcactc
     1921 attaggcacc gggatctcga ccgatgccct tgagagcctt caacccagtc agctccttcc
     1981 ggtgggcgcg gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac
     2041 tcgtaggaca ggtgccggca gcgctctggg tcattttcgg cgaggaccgc tttcgctgga
     2101 gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat cttgcacgcc ctcgctcaag
     2161 ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa gcaggccatt atcgccggca
     2221 tggcggcccc acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg ctaggctggc
     2281 ggggttgcct tactggttag cagaatgaat caccgatacg cgagcgaacg tgaagcgact
     2341 gctgctgcaa aacgtctgcg acctgagcaa caacatgaat ggtcttcggt ttccgtgttt
     2401 cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg atctgcatcg
     2461 caggatgctg ctggctaccc tgtggaacac ctacatctgt attaacgaag cgctggcatt
     2521 gaccctgagt gatttttctc tggtcccgcc gcatccatac cgccagttgt ttaccctcac
     2581 aacgttccag taaccgggca tgttcatcat cagtaacccg tatcgtgagc atcctctctc
     2641 gtttcatcgg tatcattacc cccatgaaca gaaatccccc ttacacggag gcatcagtga
     2701 ccaaacagga aaaaaccgcc cttaacatgg cccgctttat cagaagccag acattaacgc
     2761 ttctggagaa actcaacgag ctggacgcgg atgaacaggc agacatctgt gaatcgcttc
     2821 acgaccacgc tgatgagctt taccgcagct gcctcgcgcg tttcggtgat gacggtgaaa
     2881 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     2941 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga
     3001 cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat cagagcagat
     3061 tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     3121 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     3181 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     3241 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     3301 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     3361 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     3421 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     3481 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     3541 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     3601 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     3661 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     3721 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     3781 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     3841 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     3901 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     3961 taagggattt tggtcatgaa caataaaact gtctgcttac ataaacagta atacaagggg
     4021 tgttatgagc catattcaac gggaaacgtc ttgctctagg ccgcgattaa attccaacat
     4081 ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat caggtgcgac
     4141 aatctatcga ttgtatggga agcccgatgc gccagagttg tttctgaaac atggcaaagg
     4201 tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga cggaatttat
     4261 gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt tactcaccac
     4321 tgcgatcccc gggaaaacag cattccaggt attagaagaa tatcctgatt caggtgaaaa
     4381 tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg tttgtaattg
     4441 tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa tgaataacgg
     4501 tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg aacaagtctg
     4561 gaaagaaatg cataaacttt tgccattctc accggattca gtcgtcactc atggtgattt
     4621 ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg atgttggacg
     4681 agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc tcggtgagtt
     4741 ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc ctgatatgaa
     4801 taaattgcag tttcatttga tgctcgatga gtttttctaa gaattaattc atgagcggat
     4861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     4921 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     4981 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     5041 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     5101 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     5161 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     5221 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     5281 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     5341 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccaat
     5401 ccggatatag ttcctccttt cagcaaaaaa cccctcaaga cccgtttaga ggccccaagg
     5461 ggttatgcta gttattgctc agcggtggca gcagccaact cagcttcctt tcgggctttg
     5521 ttagcagccg gatctcagtg gtggtggtgg tggtgctcga gtgcggccgc aagcttcgtc
     5581 atcagatagc ctgtggc
//
