LOCUS Exported 6307 bp ds-DNA circular SYN 23-MAR-2021 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6307) AUTHORS . TITLE Direct Submission JOURNAL Exported Tuesday, Mar 23, 2021 from SnapGene 5.2.4 https://www.snapgene.com FEATURES Location/Qualifiers source 1..6307 /organism="recombinant plasmid" /mol_type="other DNA" promoter 10..54 /label=T5 promoter /note="bacteriophage T5 promoter for E. coli RNApolymerase, with embedded lac operator" protein_bind 30..46 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator toinhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." protein_bind 62..78 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator toinhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." RBS 97..108 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" misc_feature 115..192 /label=14His tag misc_feature 193..483 /gene="S. cerevisiae SMT3 (truncated)" /product="cleavable ubiquitin-like protein tag" /label=SUMO CDS 484..2499 /codon_start=1 /label=nsp14-10 fusion protein /translation="AENVTGLFKDCSKVITGLHPTQAPTHLSVDTKFKTEGLCVDIPGI PKDMTYRRLISMMGFKMNYQVNGYPNMFITREEAIRHVRAWIGFDVEGCHATREAVGTN LPLQLGFSTGVNLVAVPTGYVDTPNNTDFSRVSAKPPPGDQFKHLIPLMYKGLPWNVVR IKIVQMLSDTLKNLSDRVVFVLWAHGFELTSMKYFVKIGPERTCCLCDRRATCFSTASD TYACWHHSIGFDYVYNPFMIDVQQWGFTGNLQSNHDLYCQVHGNAHVASCDAIMTRCLA VHECFVKRVDWTIEYPIIGDELKINAACRKVQHMVVKAALLADKFPVLHDIGNPKAIKC VPQADVEWKFYDAQPCSDKAYKIEELFYSYATHSDKFTDGVCLFWNCNVDRYPANSIVC RFDTRVLSNLNLPGCDGGSLYVNKHAFHTPAFDKSAFVNLKQLPFFYYSDSPCESHGKQ VVSDIDYVPLKSATCITRCNLGGAVCRHHANEYRLYLDAYNMMISAGFSLWVYKQFDTY NLWNTFTRLQGGSGGSAGNATEVPANSTVLSFCAFAVDAAKAYKDYLASGGQPITNCVK MLCTHTGTGQAITVTPEANMDQESFGGASCCLYCRCHIDHPNPKGFCDLKGKYVQIPTT CANDPVGFTLKNTVCTVCGMWKGYGCSCDQLREPMLQ" misc_feature 484..2064 /label=coNsp14 misc_feature 2065..2082 /label=GGSGGS linker misc_feature 2083..2502 /label=coNsp10 terminator 2549..2643 /label=lambda t0 terminator /note="transcription terminator from phage lambda" CDS 2704..3519 /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" terminator 3738..3824 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnBgene" CDS complement(3918..5000) /codon_start=1 /gene="lacI" /product="lac repressor" /label=lacI /note="The lac repressor binds to the lac operator toinhibit 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" promoter complement(5001..5078) /gene="lacI (mutant)" /label=lacIq promoter /note="In the lacIq allele, a single base change in thepromoter boosts expression of the lacI gene about 10-fold." misc_feature 5270..5410 /label=bom /note="basis of mobility region from pBR322" rep_origin complement(5596..6184) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" ORIGIN 1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca 61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aaccatgagc 121 aagcatcacc atcattcagg ccatcaccat accggacacc accatcattc aggcagtcat 181 caccataccg gatcggactc agaagtcaat caagaagcta agccagaggt caagccagaa 241 gtcaagcctg agactcacat caatttaaag gtgtccgatg gatcttcaga gatcttcttc 301 aagatcaaaa agaccactcc tttaagaagg ctgatggaag cgttcgctaa aagacagggt 361 aaggaaatgg actccttaag attcttgtac gacggtatta gaattcaagc tgatcagacc 421 cctgaagatt tggacatgga ggataacgat attattgagg ctcacagaga acagattggt 481 ggcgcagaaa atgttaccgg tctgtttaaa gattgcagca aagttattac aggtctgcat 541 ccgacacagg caccgacaca tctgagcgtt gataccaaat tcaaaaccga aggtctgtgt 601 gttgatattc cgggtattcc gaaagatatg acctatcgtc gtctgattag catgatgggc 661 ttcaaaatga attatcaggt gaatggctac ccgaacatgt ttattacccg tgaagaagca 721 attcgtcacg ttcgtgcatg gattggtttt gatgttgaag gttgtcatgc aacacgtgaa 781 gcagttggca ccaatctgcc gctgcagctg ggttttagca ccggtgttaa tctggttgca 841 gttccgaccg gttatgttga taccccgaat aacaccgatt ttagccgtgt tagcgcaaaa 901 ccgcctccgg gtgatcagtt taaacatctg attccgctga tgtataaagg tctgccgtgg 961 aatgttgtgc gtatcaaaat tgttcagatg ctgagcgata ccctgaaaaa tctgagcgat 1021 cgtgttgttt ttgttctgtg ggcacatggt tttgaactga ccagcatgaa atacttcgtg 1081 aaaattggtc cggaacgtac ctgttgtctg tgtgatcgtc gtgcaacctg ttttagtacc 1141 gcaagcgata cctatgcatg ttggcatcat agcattggct tcgattatgt gtataacccg 1201 tttatgattg atgttcagca gtggggtttt accggcaatc tgcagagcaa tcatgatctg 1261 tattgtcagg ttcatggtaa tgcacatgtt gcaagctgtg atgcaattat gacccgttgt 1321 ctggcagttc atgaatgctt tgttaaacgt gttgattgga ccatcgaata tccgattatt 1381 ggtgatgagc tgaaaattaa cgcagcctgt cgtaaagttc agcacatggt ggttaaagca 1441 gcactgctgg cagataaatt tccggttctg catgatattg gtaacccgaa agcaattaaa 1501 tgtgttccgc aggcagatgt ggaatggaaa ttctatgatg cacagccgtg tagcgataaa 1561 gcctataaaa tcgaagaact gttctatagc tatgcgaccc atagcgacaa atttaccgat 1621 ggtgtttgtc tgttttggaa ttgcaatgtt gatcgctatc cggcaaatag cattgtttgc 1681 cgttttgata cccgtgttct gagcaatctg aatctgcctg gttgtgatgg tggtagcctg 1741 tatgttaaca aacatgcatt tcatacccct gcctttgata aaagcgcatt tgtgaatctg 1801 aaacagctgc cgtttttcta ttatagcgat agcccgtgtg aaagccatgg taaacaggtg 1861 gttagcgata ttgattatgt tccgctgaaa agcgcaacct gcattacccg ttgtaatctt 1921 ggtggtgcag tttgtcgtca tcatgcaaat gaatatcgtc tgtatctgga tgcctacaac 1981 atgatgatta gcgcaggttt tagcctgtgg gtgtataaac agtttgatac ctataatctg 2041 tggaacacct ttacacgtct gcagggtggt agcggtggta gtgcaggtaa tgcaaccgaa 2101 gttccggcaa atagcaccgt tctgagcttt tgtgcatttg cagttgatgc agcaaaagcc 2161 tataaagact atctggcaag cggtggtcag ccgattacca attgtgttaa aatgctgtgt 2221 acccataccg gcaccggtca ggcaattacc gttacaccgg aagcaaatat ggatcaagaa 2281 agctttggtg gtgcaagctg ttgtctgtat tgtcgttgtc atattgatca cccgaatccg 2341 aaaggttttt gtgatctgaa aggtaagtat gtgcagattc cgaccacctg tgcaaatgat 2401 ccggttggtt ttaccctgaa aaataccgtt tgcaccgttt gtggtatgtg gaaaggttat 2461 ggttgtagct gtgatcagct gcgtgaaccg atgctgcagt aataagcggc cgcacatcac 2521 catcactaag cttaattagc tgagcttgga ctcctgttga tagatccagt aatgacctca 2581 gaactccatc tggatttgtt cagaacgctc ggttgccgcc gggcgttttt tattggtgag 2641 aatccaagct agccatgaac aataaaactg tctgcttaca taaacagtaa tacaaggggt 2701 gttatgagcc atattcaacg ggaaacgtct tgctctaggc cgcgattaaa ttccaacatg 2761 gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc aggtgcgaca 2821 atctatcgat tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca tggcaaaggt 2881 agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac ggaatttatg 2941 cctcttccga ccatcaagca ttttatccgt actcctgatg atgcttggtt actcacgact 3001 gcgatccccg gcaaaacagc attccaggta ttagaagaat atcctgattc aggtgaaaat 3061 attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt ttgtaattgt 3121 ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat gaataacggt 3181 ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga acaagtctgg 3241 aaagaaatgc acaaactttt gccattctca ccggattcag tcgtcactca tggtgatttc 3301 tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga tgttggacga 3361 gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct cggtgagttt 3421 tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc tgatatgaat 3481 aaattgcagt ttcatttgat gctcgatgag tttttctaag aattaattca tgggcaaata 3541 ttatacgcaa ggcgacaagg tgctgatgcc gctggcgatt caggttcatc atgccgtttg 3601 tgatggcttc catgtcggca gaatgcttaa tgaattacaa cagtactgcg atgagtggca 3661 gggcggggcg taattttttt aaggcagtta ttggtgccct taaacgcctg gggtaatgac 3721 tctctagctt gaggcatcaa ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt 3781 ttatctgttg tttgtcggtg aacgctctcc tgagtaggac aaatccgccg ctctagatta 3841 cgtgcagtcg atgataagct gtcaaacatg agaattgtgc ctaatgagtg agctaactta 3901 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc 3961 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggggc cagggtggtt 4021 tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag 4081 agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg 4141 gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagata 4201 tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga 4261 tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt 4321 tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg 4381 cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg 4441 cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc 4501 gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga 4561 aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc 4621 ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta 4681 caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg 4741 gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg 4801 gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg 4861 taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg 4921 gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg 4981 tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat 5041 gccataccgc gaaaggtttt gcaccattcg atggtgtcgg aatttcgggc agcgttgggt 5101 cctggccacg ggtgcgcatg atctagagct gcctcgcgcg tttcggtgat gacggtgaaa 5161 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga 5221 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga 5281 cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat cagagcagat 5341 tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata 5401 ccgcatcagg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct 5461 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga 5521 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc 5581 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg 5641 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg 5701 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt 5761 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt 5821 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg 5881 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact 5941 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt 6001 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct 6061 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac 6121 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc 6181 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg 6241 ttaagggatt ttggtcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg 6301 tcttcac //