LOCUS       sequence-194699-        6064 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 6064)
  TITLE     SGC Toronto Q2 Q3 2016
REFERENCE   2  (bases 1 to 6064)
  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: 5TCM (PP-BRD3)
FEATURES             Location/Qualifiers
     source          1..6064
                     /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"
     primer_bind     complement(565..583)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      579..626
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(801..806,807..823,824..829))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(856..874)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        942..1046
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1047..1115,1116..1907)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1265..1284)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2078..2666
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2567..2586
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2820..2837
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2852..2991
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2917..2936)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3077..3099
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3093..3284)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4596..5678)
                     /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     5640..5659
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5679..5756)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5962..5981
                     /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 ctataatgag gcagatgtag ctgcgctggt gcggagcctg
      181 gatcgtgcgg aggatcatca tatctttgcc gtcgatgttc tggaaaccta tccgtactta
      241 gcggaaagct acaccaaagt atgcccgcgt cgctgcgatt tggcgaccgc agcccaaaag
      301 gcactggagg gagcttatag ctacgacctg cggttggaag gcctcaaagc cgacatcgct
      361 cttatggcgt ctaactgcgt cgcctataat gggccgacga gcgcgtatgc cgaaactgcc
      421 gccaaatttg aacggtatgc cctggaacag attgatgctt tcgtgttgga acataacggg
      481 ggttgctgat gacgaagctt gcggccgcac tcgaggatcc ggctgctaac aaagcccgaa
      541 aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct
      601 ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga tatcccgcaa
      661 gaggcccggc agtaccggca taaccaagcc tatgcctaca gcatccaggg tgacggtgcc
      721 gaggatgacg atgagcgcat tgttagattt catacacggt gcctgactgc gttagcaatt
      781 taactgtgat aaactaccgc attaaagctt atcgatgata agctgtcaaa catgagaatt
      841 cttgaagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
      901 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
      961 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     1021 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     1081 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     1141 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     1201 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     1261 ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc
     1321 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     1381 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     1441 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     1501 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     1561 caaacgacga gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat
     1621 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     1681 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     1741 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     1801 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     1861 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     1921 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     1981 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     2041 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     2101 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     2161 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     2221 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     2281 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     2341 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     2401 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     2461 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     2521 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     2581 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     2641 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     2701 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     2761 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     2821 gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc
     2881 tgtgcggtat ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata
     2941 gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac
     3001 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     3061 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     3121 cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct
     3181 gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg
     3241 ataaagcggg ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa
     3301 gggggatttc tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata
     3361 cgggttactg atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg
     3421 gtatggatgc ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat
     3481 acagatgtag gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata
     3541 atggtgcagg gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc
     3601 attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg
     3661 cgtatcggtg attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc
     3721 aacgacagga gcacgatcat gcgcacccgt ggccaggacc caacgctgcc cgagatgcgc
     3781 cgcgtgcggc tgctggagat ggcggacgcg atggatatgt tctgccaagg gttggtttgc
     3841 gcattcacag ttctccgcaa gaattgattg gctccaattc ttggagtggt gaatccgtta
     3901 gcgaggtgcc gccggcttcc attcaggtcg aggtggcccg gctccatgca ccgcgacgca
     3961 acgcggggag gcagacaagg tatagggcgg cgcctacaat ccatgccaac ccgttccatg
     4021 tgctcgccga ggcggcataa atcgccgtga cgatcagcgg tccaatgatc gaagttaggc
     4081 tggtaagagc cgcgagcgat ccttgaagct gtccctgatg gtcgtcatct acctgcctgg
     4141 acagcatggc ctgcaacgcg ggcatcccga tgccgccgga agcgagaaga atcataatgg
     4201 ggaaggccat ccagcctcgc gtcgcgaacg ccagcaagac gtagcccagc gcgtcggccg
     4261 ccatgccggc gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga
     4321 aggcttgagc gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg
     4381 cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta
     4441 cgagttgcat gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc
     4501 accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct
     4561 aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     4621 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     4681 ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc
     4741 accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga
     4801 aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg
     4861 tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt
     4921 gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc
     4981 agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct
     5041 atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc
     5101 gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga
     5161 tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc
     5221 tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg
     5281 gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga
     5341 ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg
     5401 ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc
     5461 agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt
     5521 gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt
     5581 ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg
     5641 gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc
     5701 tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg
     5761 atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag
     5821 gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc
     5881 cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg
     5941 gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt
     6001 ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg
     6061 aaat
//
