LOCUS       sequence-136207-        5753 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 5753)
  TITLE     SGC Oxford
REFERENCE   2  (bases 1 to 5753)
  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: SLX4
FEATURES             Location/Qualifiers
     source          1..5753
                     /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             610..627
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(680..698)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      694..740
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      780..1237
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(869..888)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1079..1100
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1330..2145)
                     /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(1330..2145)
                     /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     2053..2072
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2137..2156
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2267..2855
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2756..2775
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3009..3026
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3041..3183
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3109..3128)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3269..3291
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3285..3476)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4285..5367)
                     /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     5329..5348
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5368..5445)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5651..5670
                     /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 atccatgggc cgcaccttgc tctccctcgg
      181 gctgctggtt gctgactttg gcgccatggt caataaccca cacctgagtg atgtccagtt
      241 tcagacggac agcggggagg tgctttacgc ccacaagttc gtgctttatg cccgatgccc
      301 gctcctcatc cagtatgtga acaatgaagg cttctccgct atcgaggacg gggttgaaac
      361 ccagcgtgtc ctgctgggtg acgtgagcac cgaggccgcc cgcacgttcc tgcactatct
      421 ctacactgcg gacactggcc ttcctcctgg ccttagctct gagctgagct ccctggccca
      481 caggtttggc gtgagtgagc tcgttcacct gtgcgaacag gtgcctattg ccactgactc
      541 agagtgacag taaaggtgga tacggatccg aattcgagct ccgtcgacaa gcttgcggcc
      601 gcactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa
      661 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
      721 cgggtcttga ggggtttttt tgctgaaaag gaggaactat atccggattg gcgaatggga
      781 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
      841 tacactttgc cagcgcccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
      901 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
      961 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     1021 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     1081 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     1141 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     1201 aacgcgaatt ttaacaaaat attaacgttt acaatttcag gtggcacttt tcggggaaat
     1261 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     1321 aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg
     1381 attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag
     1441 gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc
     1501 aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg
     1561 agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc agacttgttc
     1621 aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat
     1681 tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac
     1741 aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga
     1801 atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag tggtgagtaa
     1861 ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt
     1921 cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg
     1981 tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg tcgcacctga
     2041 ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt
     2101 taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt
     2161 actgtttatg taagcagaca gttttattgt tcatgaccaa aatcccttaa cgtgagtttt
     2221 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     2281 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     2341 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     2401 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     2461 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     2521 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     2581 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     2641 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     2701 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     2761 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     2821 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     2881 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     2941 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     3001 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc
     3061 ttacgcatct gtgcggtatt tcacaccgca tatatggtgc actctcagta caatctgctc
     3121 tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct
     3181 gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     3241 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     3301 tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat
     3361 tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat
     3421 gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat
     3481 gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg
     3541 atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt
     3601 aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag
     3661 cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag
     3721 atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg
     3781 aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt
     3841 cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct
     3901 agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc catgccggcg
     3961 ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa ggcttgagcg
     4021 agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc gctccagcga
     4081 aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac gagttgcatg
     4141 ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca ccggaaggag
     4201 ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta atgagtgagc
     4261 taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     4321 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag
     4381 ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc
     4441 ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt
     4501 gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac
     4561 cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc
     4621 catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat
     4681 ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat
     4741 ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact
     4801 taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc
     4861 cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac
     4921 atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc
     4981 atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc
     5041 cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag
     5101 ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact
     5161 ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt
     5221 gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac
     5281 gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc
     5341 gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg
     5401 ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct
     5461 ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg ccgttgagca
     5521 ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc ccggccacgg
     5581 ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat
     5641 cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg gcgccggtga
     5701 tgccggccac gatgcgtccg gcgtagagga tcgagatctc gatcccgcga aat
//
