LOCUS Exported 5114 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Used for expression of protein in E. coli as TEV cleavable N-terminual 8His and DsbC (leaderless thiol:disulfide interchange protein DsbC from E. coli) fusions with optional C-terminal StrepII-tag. ACCESSION . VERSION . KEYWORDS pExp-DsbC SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5114) AUTHORS Lulla A, Hyvonen M TITLE Protein expressionists toolkit JOURNAL unpublished REFERENCE 2 (bases 1 to 5114) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5114 /organism="synthetic DNA construct" /mol_type="other DNA" 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 831..854 /codon_start=1 /product="peptide that binds Strep-Tactin(R), an engineered form of streptavidin" /label=Strep-Tag II /translation="WSHPQFEK" promoter complement(868..886) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(869..886) /label=SP6 /note="SP6 promoter, forward primer" primer_bind complement(944..962) /label=T7 Term /note="T7 terminator, reverse primer" terminator 958..1005 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" primer_bind complement(1241..1262) /label=F1ori-F /note="F1 origin, forward primer" CDS complement(1412..2494) /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 2456..2475 /label=LacI-R /note="LacI, reverse primer" promoter complement(2495..2572) /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 2711..2730 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind complement(2884..2903) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 3003..3025 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(3063..3081) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 3149..3253 /gene="bla" /label=AmpR promoter CDS 3254..4114 /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(3472..3491) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4285..4873 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4774..4793 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5027..5044 /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 ctgtggttac tgccacaaac 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 tggtaccgaa aacctgtact tccagtgaga ccttaattaa ctcgagcgca tggagccacc 841 cgcagttcga aaaataagct tgagtattct atagtgtcac ctaaatccca gcttgatccg 901 gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga gcaataacta 961 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa aggaggaact 1021 atatccggat aacctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt 1081 tgcgcagcct gaatggcgaa tggaaattgt aagcgttaat attttgttaa aattcgcgtt 1141 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta 1201 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc 1261 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg 1321 cccacagctt gcatgcctgc aggtcggaag cataaagtgt aaagcctggg gtgcctaatg 1381 agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct 1441 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg 1501 gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg 1561 cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat 1621 cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc 1681 ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc 1741 ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca 1801 tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg 1861 gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga 1921 cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct 1981 ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt 2041 cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat 2101 cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt 2161 gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg 2221 cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg 2281 ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca 2341 cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg 2401 cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat 2461 actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt 2521 ccgggcgcta tcatgccata ccgcgaaagg ttttgcacca ttcgatggtg tcggcgagct 2581 cgaattgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa 2641 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg 2701 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg 2761 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcct 2821 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatat ggtgcactct 2881 cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc 2941 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt 3001 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa 3061 gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac 3121 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat 3181 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg 3241 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc 3301 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga 3361 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga 3421 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg 3481 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc 3541 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac 3601 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact 3661 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca 3721 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg 3781 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact 3841 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg 3901 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg 3961 tgagcgtggt tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat 4021 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc 4081 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat 4141 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt 4201 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc 4261 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt 4321 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac 4381 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt 4441 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct 4501 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga 4561 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac 4621 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg 4681 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt 4741 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc 4801 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg 4861 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc 4921 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc 4981 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag 5041 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca 5101 ttaatgcagc tgat //