LOCUS       sequence-194479-        6915 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 6915)
  TITLE     SGC Oxford
REFERENCE   2  (bases 1 to 6915)
  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: ASB11
FEATURES             Location/Qualifiers
     source          1..6915
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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             92..109
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             134..154
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     promoter        894..912
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    913..937
                     /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             952..974
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     promoter        1396..1414
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1415..1439
                     /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             1454..1476
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1780..1797
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1850..1868)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1864..1911
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1948..2403
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2035..2054)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2245..2266
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2495..3310)
                     /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(2495..3310)
                     /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     3218..3237
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3302..3321
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3432..4020
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3921..3940
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4174..4191
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4206..4345
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4271..4290)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4431..4453
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4447..4638)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5447..6529)
                     /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     6491..6510
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6530..6607)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6813..6832
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gcaccatcat catcatcatt cttctggtgt
      121 agatctgggt accgagaacc tgtacttcca atccatggct gatcgatccc cacttcatga
      181 agctgcagct caggggcgct tactggccct taaaacttta attgcacaag gtgtcaatgt
      241 gaaccttgtg acaattaacc gggtgtcttc tctccacgag gcatgccttg gaggtcacgt
      301 ggcctgtgcc aaagccttat tggaaaatgg tgcacacgtc aatggagtga cagttcacgg
      361 agccacaccc ctcttcaatg cttgctgcag cggcagtgct gcatgtgtca atgtgctgct
      421 ggagttcgga gccaaggccc agttggaggt gcacctggcc tcgcccatcc atgaggcagt
      481 gaagagaggt cacagagagt gcatggagat cctgctggca aataatgtta acattgacca
      541 tgaggtgcct cagctcggaa ctcccctata tgtggcctgc acctaccaga gggtagactg
      601 tgtgaagaaa cttctagaat taggagccag tgtcgaccat ggccagtggc tggacacccc
      661 actccatgct gcagcgaggc agtccaatgt ggaggtcatc cacctgctaa ccgactatgg
      721 agctaacctg aagcgtagaa atgctcaggg caaaagtgcg cttgatctgg cggctccaaa
      781 aagcagcgtg gagcaggcac tcttgctccg tgaaggccca cctgctcttt gacagtaaag
      841 gtggatacgg atcccccctt ggggcctcta aacgggtctt gaggggtttt ttgtaatacg
      901 actcactata ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa
      961 ctttaagaag gagatatacc atggacgtgt tcctcatgat ccggcgccac aagaccacca
     1021 tcttcacgga cgccaaggag tccagcacgg tgttcgaact gaagcgcatc gtcgagggca
     1081 tcctcaagcg gcctcctgac gagcagcggc tgtacaagga tgaccaactc ttggatgatg
     1141 gcaagacact gggcgagtgt ggcttcacca gtcaaacagc acggccacag gccccagcca
     1201 cagtggggct ggccttccgg gcagatgaca ccttcgaggc cctgtgcatc gagccgtttt
     1261 ccagcccgcc agagctgccc gatgtgatga agccccagga ctcgggaagc agtgccaatg
     1321 aacaagccgt gcagtgagag ctccgtcgac aagcttcccc ttggggcctc taaacgggtc
     1381 ttgaggggtt ttttgtaata cgactcacta taggggaatt gtgagcggat aacaattccc
     1441 ctctagaaat aattttgttt aactttaaga aggagatata ccatgtatgt caaattgata
     1501 tcatctgatg gccatgaatt tattgtaaaa agagaacatg cattaacatc aggcacgata
     1561 aaagccatgt tgagtggccc aggtcagttt gctgagaacg aaaccaatga ggtcaatttt
     1621 agagagatac cttcacatgt gctatcgaaa gtatgcatgt attttacgta caaggttcgc
     1681 tacactaaca gctccaccga gattcctgaa ttcccaattg cacctgaaat tgcactggaa
     1741 ctgctgatgg ctgcgaactt cttagattgt taactcgagc accaccacca ccaccactga
     1801 gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa
     1861 taactagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaagga
     1921 ggaactatat ccggattggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
     1981 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     2041 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     2101 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     2161 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     2221 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     2281 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     2341 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     2401 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     2461 acattcaaat atgtatccgc tcatgaatta attcttagaa aaactcatcg agcatcaaat
     2521 gaaactgcaa tttattcata tcaggattat caataccata tttttgaaaa agccgtttct
     2581 gtaatgaagg agaaaactca ccgaggcagt tccataggat ggcaagatcc tggtatcggt
     2641 ctgcgattcc gactcgtcca acatcaatac aacctattaa tttcccctcg tcaaaaataa
     2701 ggttatcaag tgagaaatca ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt
     2761 tatgcatttc tttccagact tgttcaacag gccagccatt acgctcgtca tcaaaatcac
     2821 tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg agcgagacga aatacgcgat
     2881 cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca
     2941 gcgcatcaac aatattttca cctgaatcag gatattcttc taatacctgg aatgctgttt
     3001 tcccggggat cgcagtggtg agtaaccatg catcatcagg agtacggata aaatgcttga
     3061 tggtcggaag aggcataaat tccgtcagcc agtttagtct gaccatctca tctgtaacat
     3121 cattggcaac gctacctttg ccatgtttca gaaacaactc tggcgcatcg ggcttcccat
     3181 acaatcgata gattgtcgca cctgattgcc cgacattatc gcgagcccat ttatacccat
     3241 ataaatcagc atccatgttg gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa
     3301 tatggctcat aacacccctt gtattactgt ttatgtaagc agacagtttt attgttcatg
     3361 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     3421 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     3481 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     3541 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     3601 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     3661 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     3721 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     3781 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     3841 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     3901 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3961 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     4021 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     4081 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     4141 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     4201 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaatggt
     4261 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc
     4321 gctacgtgac tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg
     4381 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     4441 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc
     4501 atcagcgtgg tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt
     4561 gagtttctcc agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt
     4621 tttttcctgt ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat
     4681 gataccgatg aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg
     4741 gttactggaa cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa
     4801 aatcactcag ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag
     4861 ccagcagcat cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt
     4921 ttccagactt tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga
     4981 cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc
     5041 agtaaggcaa ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac
     5101 ccgtggggcc gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg
     5161 accagtgacg aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc
     5221 gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg
     5281 cacctgtcct acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat
     5341 gccccgcgcc caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga
     5401 tcccggtgcc taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt
     5461 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     5521 cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct
     5581 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc
     5641 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt
     5701 cggtatcgtc gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa
     5761 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga
     5821 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt
     5881 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac
     5941 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca
     6001 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt
     6061 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt
     6121 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt
     6181 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg
     6241 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg
     6301 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg
     6361 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt
     6421 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat
     6481 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc
     6541 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga
     6601 tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt
     6661 agtaggttga ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg
     6721 cccaacagtc ccccggccac ggggcctgcc accataccca cgccgaaaca agcgctcatg
     6781 agcccgaagt ggcgagcccg atcttcccca tcggtgatgt cggcgatata ggcgccagca
     6841 accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag gatcgagatc
     6901 tcgatcccgc gaaat
//
