LOCUS       sequence-247823-        5126 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5126)
  AUTHORS   Lulla A, Hyvonen M
  TITLE     Protein expressionists toolkit
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 5126)
  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"; journalName: 
            "unpublished"
COMMENT     Sequence Label: pExp-DsbC(SS)
FEATURES             Location/Qualifiers
     source          1..5126
                     /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    19..43
                     /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             56..78
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             92..115
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             843..866
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     promoter        complement(880..898)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(881..898)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(956..974)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      970..1017
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1253..1274)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1424..2506)
                     /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     2468..2487
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2507..2584)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     2723..2742
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(2896..2915)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3015..3037
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3075..3093)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3161..3265
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3266..3334,3335..4126)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3484..3503)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4297..4885
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4786..4805
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5039..5056
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatacgact cactataggg aattgtgagc ggataacaat tcctctagaa ataattttgt
       61 ttaactttaa gaaggagata tatccatgaa tcaccatcac catcaccatc accattctgg
      121 cactagtggc gatgacgcgg caattcaaca aacgttagcc aaaatgggca tcaaaagcag
      181 cgatattcag cccgcgcctg tagctggcat gaagacagtt ctgactaaca gcggcgtgtt
      241 gtacatcacc gatgatggta aacatatcat tcaggggcca atgtatgacg ttagtggcac
      301 ggctccggtc aatgtcacca ataagatgct gttaaagcag ttgaatgcgc ttgaaaaaga
      361 gatgatcgtt tataaagcgc cgcaggaaaa acacgtcatc accgtgttta ctgatattac
      421 ctctggttac tcccacaaac tgcatgagca aatggcagac tacaacgcgc tggggatcac
      481 cgtgcgttat cttgctttcc cgcgccaggg gctggacagc gatgcagaga aagaaatgaa
      541 agctatctgg tgtgcgaaag ataaaaacaa agcgtttgat gatgtgatgg caggtaaaag
      601 cgtcgcacca gccagttgcg acgtggatat tgccgaccat tacgcacttg gcgtccagct
      661 tggcgttagc ggtactccgg cagttgtgct gagcaatggc acacttgttc cgggttacca
      721 gccgccgaaa gagatgaaag aattcctcga cgaacaccaa aaaatgacca gcggtaaatc
      781 tggtacctcc ggttctactg aaaacctgta cttccagtga gaccttaatt aactcgagcg
      841 catggagcca cccgcagttc gaaaaataag cttgagtatt ctatagtgtc acctaaatcc
      901 cagcttgatc cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct
      961 gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg
     1021 aaaggaggaa ctatatccgg ataacctggc gtaatagcga agaggcccgc accgatcgcc
     1081 cttcccaaca gttgcgcagc ctgaatggcg aatggaaatt gtaagcgtta atattttgtt
     1141 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     1201 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     1261 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     1321 tcagggcgat ggcccacagc ttgcatgcct gcaggtcgga agcataaagt gtaaagcctg
     1381 gggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca
     1441 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     1501 tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat
     1561 tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca
     1621 gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag ctgtcttcgg
     1681 tatcgtcgta tcccactacc gagatatccg caccaacgcg cagcccggac tcggtaatgg
     1741 cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc
     1801 cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc
     1861 gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca
     1921 gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg
     1981 cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga
     2041 tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca
     2101 cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg
     2161 cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca
     2221 ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg
     2281 gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca
     2341 gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt
     2401 cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag
     2461 agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc accaccctga
     2521 attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcac cattcgatgg
     2581 tgtcggcgag ctcgaattgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     2641 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     2701 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     2761 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     2821 agggcgcgtc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     2881 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     2941 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     3001 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     3061 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     3121 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     3181 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     3241 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     3301 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     3361 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     3421 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     3481 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     3541 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     3601 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     3661 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     3721 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     3781 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     3841 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     3901 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     3961 atctggagcc ggtgagcgtg gttctcgcgg tatcattgca gcactggggc cagatggtaa
     4021 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     4081 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     4141 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     4201 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     4261 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     4321 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     4381 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     4441 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     4501 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     4561 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     4621 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     4681 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     4741 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     4801 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     4861 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     4921 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     4981 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     5041 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
     5101 ttggccgatt cattaatgca gctgat
//
