LOCUS       sequence-193497-        5774 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 5774)
  TITLE     SGC Toronto Q4 2016, Q1 Q2 2017
REFERENCE   2  (bases 1 to 5774)
  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: 5VS7 (PP-BRD4)
FEATURES             Location/Qualifiers
     source          1..5774
                     /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             91..108
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             121..141
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             525..542
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(595..613)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      609..656
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      693..1148
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(780..799)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     990..1011
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1240..2055)
                     /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(1240..2055)
                     /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     1963..1982
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2047..2066
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2177..2765
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2666..2685
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2919..2936
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2951..3090
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3016..3035)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3176..3198
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3192..3383)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     5350..5369
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5389..5466)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5672..5691
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg catcatcatc atcatcacag cagcggcaga
      121 gaaaacttgt atttccaggg ctatgacgag attgaagaat taaagagcaa gaatgaagta
      181 ctaacaaatt tattaaataa gctaatagct tttgataaga aaagaatatt tttatatcct
      241 gtgaatgtac agttagtacc tgattatttg aatgtcataa aagaacctat ggattttaca
      301 acaatgaaac agaaacttca gaattttaaa tataaaagtt ttcaagaatt tgagaaagat
      361 gttcttttaa ttattaataa ttgttataca tataatgacc caagtacaat atattataaa
      421 tttgctgaag atatagaaac ttattataaa aaattaaata ttaaaattca aacgaaatat
      481 atgaacatac acctatgatg acgaagcttg cggccgcact cgagcaccac caccaccacc
      541 actgagatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct gccaccgctg
      601 agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgctga
      661 aaggaggaac tatatccgga ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc
      721 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
      781 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
      841 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
      901 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
      961 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     1021 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     1081 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
     1141 tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     1201 taaatacatt caaatatgta tccgctcatg aattaattct tagaaaaact catcgagcat
     1261 caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg
     1321 tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta
     1381 tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa
     1441 aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa
     1501 aagtttatgc atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa
     1561 atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac
     1621 gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac
     1681 tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc
     1741 tgttttcccg gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg
     1801 cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt
     1861 aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt
     1921 cccatacaat cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata
     1981 cccatataaa tcagcatcca tgttggaatt taatcgcggc ctagagcaag acgtttcccg
     2041 ttgaatatgg ctcataacac cccttgtatt actgtttatg taagcagaca gttttattgt
     2101 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     2161 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     2221 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     2281 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt
     2341 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     2401 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     2461 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     2521 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     2581 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     2641 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     2701 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     2761 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     2821 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     2881 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     2941 cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
     3001 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg
     3061 ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg
     3121 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     3181 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa
     3241 agctcatcag cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc
     3301 tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg
     3361 gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg
     3421 gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat
     3481 gcccggttac tggaacgttg tgagggtaaa caactggcgg tatggatgcg gcgggaccag
     3541 agaaaaatca ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag
     3601 ggtagccagc agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc
     3661 cgcgtttcca gactttacga aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc
     3721 gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc
     3781 taaccagtaa ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg
     3841 cgcacccgtg gggccgccat gccggcgata atggcctgct tctcgccgaa acgtttggtg
     3901 gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac cgcaagcgac
     3961 aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac ccagagcgct
     4021 gccggcacct gtcctacgag ttgcatgata aagaagacag tcataagtgc ggcgacgata
     4081 gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa gggcatcggt
     4141 cgagatcccg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc
     4201 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     4261 agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa
     4321 cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggc
     4381 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
     4441 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
     4501 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
     4561 ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat
     4621 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
     4681 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
     4741 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
     4801 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
     4861 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
     4921 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
     4981 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
     5041 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
     5101 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
     5161 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
     5221 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
     5281 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
     5341 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
     5401 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
     5461 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
     5521 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc
     5581 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
     5641 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
     5701 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct
     5761 cgatcccgcg aaat
//
