LOCUS       sequence-136224-        5857 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 5857)
  TITLE     SGC Oxford
REFERENCE   2  (bases 1 to 5857)
  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: SHC1
FEATURES             Location/Qualifiers
     source          1..5857
                     /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             718..735
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(788..806)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      802..849
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      886..1341
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(973..992)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1183..1204
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1434..2249)
                     /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(1434..2249)
                     /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     2157..2176
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2241..2260
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2371..2959
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2860..2879
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3113..3130
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3145..3287
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3213..3232)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3373..3395
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3389..3580)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4389..5471)
                     /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     5433..5452
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5472..5549)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5755..5774
                     /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 tggctgcatc ccaacgacaa
      181 agtcatggga cccggggttt cctacttggt tcggtacatg ggttgtgtgg aggtcctcca
      241 gtcaatgcgt gccctggact tcaacacccg gactcaggtc accagggagg ccatcagtct
      301 ggtgtgtgag gctgtgccgg gtgctaaggg ggcgacaagg aggagaaagc cctgtagccg
      361 cccgctcagc tctatcctgg ggaggagtaa cctgaaattt gctggaatgc caatcactct
      421 caccgtctcc accagcagcc tcaacctcat ggccgcagac tgcaaacaga tcatcgccaa
      481 ccaccacatg caatctatct catttgcatc cggcggggat ccggacacag ccgagtatgt
      541 cgcctatgtt gccaaagacc ctgtgaatca gagagcctgc cacattctgg agtgtcccga
      601 agggcttgcc caggatgtca tcagcaccat tggccaggcc ttcgagttgc gctgacagta
      661 aaggtggata cggatccgaa ttcgagctcc gtcgacaagc ttgcggccgc actcgagcac
      721 caccaccacc accactgaga tccggctgct aacaaagccc gaaaggaagc tgagttggct
      781 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
      841 ggttttttgc tgaaaggagg aactatatcc ggattggcga atgggacgcg ccctgtagcg
      901 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
      961 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     1021 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     1081 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
     1141 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     1201 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     1261 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     1321 aaatattaac gtttacaatt tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     1381 tttgtttatt tttctaaata cattcaaata tgtatccgct catgaattaa ttcttagaaa
     1441 aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat
     1501 ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg
     1561 gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat
     1621 ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc
     1681 ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta
     1741 cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga
     1801 gcgagacgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac
     1861 cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct
     1921 aatacctgga atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga
     1981 gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg
     2041 accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct
     2101 ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg
     2161 cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag
     2221 caagacgttt cccgttgaat atggctcata acaccccttg tattactgtt tatgtaagca
     2281 gacagtttta ttgttcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     2341 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     2401 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     2461 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct
     2521 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     2581 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     2641 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     2701 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     2761 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     2821 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     2881 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     2941 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     3001 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     3061 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     3121 agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg
     3181 tatttcacac cgcatatatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3241 agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc
     3301 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     3361 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     3421 cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct
     3481 gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa
     3541 agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg
     3601 gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg
     3661 ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat
     3721 ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag
     3781 atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg
     3841 tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc
     3901 atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta
     3961 tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg
     4021 acaggagcac gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg gcctgcttct
     4081 cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc
     4141 cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga
     4201 aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca
     4261 taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg
     4321 ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg
     4381 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4441 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc
     4501 accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc
     4561 aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc
     4621 gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca
     4681 acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca
     4741 accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg
     4801 gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga
     4861 tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac
     4921 agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct
     4981 tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc
     5041 ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta
     5101 atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg
     5161 acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat
     5221 ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca
     5281 atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc
     5341 tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc
     5401 accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt
     5461 actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg
     5521 cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc
     5581 ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa
     5641 tggtgcatgc aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc
     5701 cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc cgatcttccc catcggtgat
     5761 gtcggcgata taggcgccag caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg
     5821 tccggcgtag aggatcgaga tctcgatccc gcgaaat
//
