LOCUS       sequence-319827-        5642 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 5642)
  TITLE     SGC Toronto 2019-Q1 to 2020-Q2
REFERENCE   2  (bases 1 to 5642)
  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: 6OQM
FEATURES             Location/Qualifiers
     source          1..5642
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             366..383
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(436..454)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      450..497
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      534..989
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(621..640)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     831..852
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1081..1896)
                     /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(1081..1896)
                     /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     1804..1823
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1888..1907
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2018..2606
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2507..2526
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2760..2777
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2792..2931
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2857..2876)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3017..3039
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3033..3224)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4033..5115)
                     /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     5077..5096
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5116..5193)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5399..5418
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5502..5521
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5502..5520
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5521..5545
                     /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             5560..5582
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5592..5609
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5622..5642
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
ORIGIN
        1 agttgtgact tctcaccggg agatttggtt tgggccaaga tggagggtta cccctggtgg
       61 ccttgtctgg tttacaacca cccctttgat ggaacattca tccgcgagaa agggaaatca
      121 gtccgtgttc atgtacagtt ttttgatgac agcccaacaa ggggctgggt tagcaaaagg
      181 cttttaaagc catatacagg ttcaaaatca aaggaagccc agaagggagg tcatttttac
      241 agtgcaaagc ctgaaatact gagagcaatg caacgtgcag acgaagcctt aaataaagac
      301 aagattaaga ggcttgaatt ggcagtttgt gatgagtgat gacgaagctt gcggccgcac
      361 tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga aaggaagctg
      421 agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
      481 tcttgagggg ttttttgctg aaaggaggaa ctatatccgg attggcgaat gggacgcgcc
      541 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
      601 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
      661 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
      721 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
      781 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
      841 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
      901 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
      961 ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga
     1021 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gaattaattc
     1081 ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat
     1141 accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca
     1201 taggatggca agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc
     1261 tattaatttc ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac
     1321 tgaatccggt gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca
     1381 gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg
     1441 cgcctgagcg agacgaaata cgcgatcgct gttaaaagga caattacaaa caggaatcga
     1501 atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata
     1561 ttcttctaat acctggaatg ctgttttccc ggggatcgca gtggtgagta accatgcatc
     1621 atcaggagta cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt
     1681 tagtctgacc atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa
     1741 caactctggc gcatcgggct tcccatacaa tcgatagatt gtcgcacctg attgcccgac
     1801 attatcgcga gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg
     1861 cctagagcaa gacgtttccc gttgaatatg gctcataaca ccccttgtat tactgtttat
     1921 gtaagcagac agttttattg ttcatgacca aaatccctta acgtgagttt tcgttccact
     1981 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     2041 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     2101 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     2161 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     2221 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     2281 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     2341 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     2401 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     2461 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     2521 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     2581 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     2641 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     2701 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     2761 gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc
     2821 tgtgcggtat ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata
     2881 gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac
     2941 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     3001 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     3061 cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct
     3121 gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg
     3181 ataaagcggg ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa
     3241 gggggatttc tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata
     3301 cgggttactg atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg
     3361 gtatggatgc ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat
     3421 acagatgtag gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata
     3481 atggtgcagg gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc
     3541 attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg
     3601 cgtatcggtg attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc
     3661 aacgacagga gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc
     3721 ttctcgccga aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag
     3781 attccgaata ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg
     3841 ccgaaaatga cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca
     3901 gtcataagtg cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg
     3961 aaggctctca agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta
     4021 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     4081 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct
     4141 tttcaccagt gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg
     4201 cagcaagcgg tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa
     4261 cggcgggata taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc
     4321 accaacgcgc agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt
     4381 ggcaaccagc atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa
     4441 accggacatg gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt
     4501 gagatattta tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc
     4561 taacagcgcg atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc
     4621 gtcttcatgg gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa
     4681 cgccggaaca ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata
     4741 gttaatgatc agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc
     4801 ttcgacgccg cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg
     4861 agatttaatc gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac
     4921 gccaatcagc aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt
     4981 cagctccgcc atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg
     5041 gttcaccacg cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa
     5101 cgttactggt ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat
     5161 accgcgaaag gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg
     5221 actcctgcat taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa
     5281 ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca
     5341 tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg
     5401 tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga
     5461 tgcgtccggc gtagaggatc gagatctcga tcccgcgaaa ttaatacgac tcactatagg
     5521 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga
     5581 gatataccat gcatcatcat catcatcaca gcagcggcag agaaaacttg tatttccagg
     5641 gc
//
