LOCUS       sequence-234360-        6402 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression for structure determination.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6402)
  TITLE     SGC Toronto Q3-Q4 2017, Q1-Q2 2018
REFERENCE   2  (bases 1 to 6402)
  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: 6CUI (BRD2)
FEATURES             Location/Qualifiers
     source          1..6402
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             12..34
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             42..695
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     regulatory      720..729
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             822..842
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1220..1237
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1290..1308)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1304..1351
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1388..1843
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1475..1494)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1685..1706
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1935..2750)
                     /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(1935..2750)
                     /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     2658..2677
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2742..2761
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2872..3460
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3361..3380
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3614..3631
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3646..3785
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3711..3730)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3871..3893
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3887..4078)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4887..5969)
                     /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     5931..5950
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5970..6047)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6253..6272
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6356..6375
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6356..6374
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6375..6399
                     /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)."
ORIGIN
        1 ctagaaataa ttttgtttaa ctttaagaag gagatataca tatgtcccct atactaggtt
       61 attggaaaat taagggcctt gtgcaaccca ctcgacttct tttggaatat cttgaagaaa
      121 aatatgaaga gcatttgtat gagcgcgatg aaggtgataa atggcgaaac aaaaagtttg
      181 aattgggttt ggagtttccc aatcttcctt attatattga tggtgatgtt aaattaacac
      241 agtctatggc catcatacgt tatatagctg acaagcacaa catgttgggt ggttgtccaa
      301 aagagcgtgc agagatttca atgcttgaag gagcggtttt ggatattaga tacggtgttt
      361 cgagaattgc atatagtaaa gactttgaaa ctctcaaagt tgattttctt agcaagctac
      421 ctgaaatgct gaaaatgttc gaagatcgtt tatgtcataa aacatattta aatggtgatc
      481 atgtaaccca tcctgacttc atgttgtatg acgctcttga tgttgtttta tacatggacc
      541 caatgtgcct ggatgcgttc ccaaaattag tttgttttaa aaaacgtatt gaagctatcc
      601 cacaaattga taagtacttg aaatccagca agtatatagc atggcctttg cagggctggc
      661 aagccacgtt tggtggtggc gaccatcctc caaaatcgga tggttcatcc gaaggagacg
      721 ccaccatggg gcatatggtg cacaggcctt tccaatccgc ggccagttac cccaaccggc
      781 ccagcaagac acccccccga gctgcagaga gcagcggcag agaaaacttg tatttccagg
      841 gcgaggagga agagagcagg cccatgagtt acgatgagaa gcggcagctg agcctggaca
      901 tcaacaaatt acctggggag aagctgggcc gagttgtgca tataatccaa gccagggagc
      961 cctctttacg tgattcaaac ccagaagaga ttgagattga ttttgaaaca ctcaagccat
     1021 ccacacttag agagcttgag cgctatgtcc tttcctgcct acgtaagaaa ccccggaagc
     1081 cctacaccat taagaagcct gtgggaaaga caaaggagga actggctttg gagaaaaagc
     1141 gggaattaga aaagcggtta caagatgtca gcggacagct caattctact tgatgacgaa
     1201 gcttgcggcc gcactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc
     1261 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     1321 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
     1381 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1441 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     1501 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     1561 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     1621 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     1681 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     1741 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     1801 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     1861 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     1921 tcatgaatta attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata
     1981 tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca
     2041 ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca
     2101 acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca
     2161 ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact
     2221 tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta
     2281 ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta
     2341 caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca
     2401 cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg
     2461 agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat
     2521 tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg
     2581 ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca
     2641 cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg
     2701 gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt
     2761 gtattactgt ttatgtaagc agacagtttt attgttcatg accaaaatcc cttaacgtga
     2821 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     2881 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     2941 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     3001 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     3061 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     3121 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     3181 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     3241 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     3301 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     3361 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     3421 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     3481 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     3541 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     3601 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
     3661 tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag tacaatctgc
     3721 tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg
     3781 ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     3841 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     3901 cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg
     3961 attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta
     4021 atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt ttggtcactg
     4081 atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg aaacgagaga
     4141 ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa cgttgtgagg
     4201 gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag ggtcaatgcc
     4261 agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat cctgcgatgc
     4321 agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt tacgaaacac
     4381 ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc
     4441 ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa ccccgccagc
     4501 ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggggcc gccatgccgg
     4561 cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag
     4621 cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc
     4681 gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca
     4741 tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg
     4801 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
     4861 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     4921 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
     4981 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
     5041 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
     5101 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
     5161 accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
     5221 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
     5281 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
     5341 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
     5401 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
     5461 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
     5521 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
     5581 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
     5641 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
     5701 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
     5761 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
     5821 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
     5881 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
     5941 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
     6001 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
     6061 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     6121 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggccac
     6181 ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg
     6241 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
     6301 gatgccggcc acgatgcgtc cggcgtagag gatcgagatc tcgatcccgc gaaattaata
     6361 cgactcacta taggggaatt gtgagcggat aacaattccc ct
//
