LOCUS       sequence-150431-        5758 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 5758)
  TITLE     SGC Oxford
REFERENCE   2  (bases 1 to 5758)
  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: CBS
FEATURES             Location/Qualifiers
     source          1..5758
                     /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             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"
     CDS             619..636
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(689..707)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      703..750
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      787..1242
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(874..893)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1084..1105
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1335..2150)
                     /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(1335..2150)
                     /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     2058..2077
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2142..2161
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2272..2860
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2761..2780
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3014..3031
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3046..3188
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3114..3133)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3274..3296
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3290..3481)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4290..5372)
                     /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     5334..5353
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5373..5450)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5656..5675
                     /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 atccatgaag ccctggtggt ggcacctccg
      181 tgttcaggag ctgggcctgt cagccccgct gaccgtgctc ccgaccatca cctgtgggca
      241 caccatcgag atcctccggg agaagggctt cgaccaggcg cccgtggtgg atgaggcggg
      301 ggtaatcctg ggaatggtga cgcttgggaa catgctctcg tccctgcttg ccgggaaggt
      361 gcagccgtca gaccaagttg gcaaagtcat ctacaagcag ttcaaacaga tccgcctcac
      421 ggacacgctg ggcaggctct cgcacatcct ggagatggac cacttcgccc tggtggtgca
      481 cgagcagcag cggcagatgg tgttcggggt ggtcaccgcc attgacttgc tgaacttcgt
      541 ggccgcccag gagtgacagt aaaggtggat acggatccga attcgagctc cgtcgacaag
      601 cttgcggccg cactcgagca ccaccaccac caccactgag atccggctgc taacaaagcc
      661 cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg
      721 gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc cggattggcg
      781 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
      841 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
      901 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
      961 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     1021 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     1081 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     1141 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     1201 aatttaacgc gaattttaac aaaatattaa cgtttacaat ttcaggtggc acttttcggg
     1261 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     1321 tcatgaatta attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata
     1381 tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca
     1441 ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca
     1501 acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca
     1561 ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact
     1621 tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta
     1681 ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta
     1741 caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca
     1801 cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg
     1861 agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat
     1921 tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg
     1981 ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca
     2041 cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg
     2101 gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt
     2161 gtattactgt ttatgtaagc agacagtttt attgttcatg accaaaatcc cttaacgtga
     2221 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     2281 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     2341 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     2401 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     2461 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     2521 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     2581 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     2641 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     2701 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     2761 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     2821 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     2881 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     2941 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     3001 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
     3061 tctccttacg catctgtgcg gtatttcaca ccgcatatat ggtgcactct cagtacaatc
     3121 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     3181 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     3241 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     3301 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     3361 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     3421 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     3481 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     3541 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     3601 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     3661 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     3721 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     3781 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     3841 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     3901 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc
     3961 cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt
     4021 gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc
     4081 agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt
     4141 gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga
     4201 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     4261 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4321 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4381 gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc
     4441 tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc
     4501 tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc
     4561 actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc
     4621 agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt
     4681 tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc
     4741 tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca
     4801 gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc
     4861 acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca
     4921 gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc
     4981 tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc
     5041 accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca
     5101 cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc
     5161 agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg
     5221 cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca
     5281 gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac
     5341 tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc
     5401 gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg
     5461 acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt
     5521 gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc
     5581 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
     5641 ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc
     5701 ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc cgcgaaat
//
