LOCUS Exported 6273 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expresses SpyCatcher003-mi3 in bacterial cells as virus-like particle platform for plug-and-display vaccination. ACCESSION . VERSION . KEYWORDS SpyCatcher003-mi3 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6273) AUTHORS Rahikainen R, Rijal P, Tan TK, Wu HJ, Andersson AC, Barrett JR, Bowden TA, Draper SJ, Townsend AR, Howarth M TITLE Overcoming Symmetry Mismatch in Vaccine Nanoassembly through Spontaneous Amidation. JOURNAL Angew Chem Int Ed Engl. 2020 Sep 4. doi: 10.1002/anie.202009663. PUBMED 32886840 REFERENCE 2 (bases 1 to 6273) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6273 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 26..73 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" primer_bind 69..87 /label=T7 Term /note="T7 terminator, reverse primer" CDS complement(140..157) /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" RBS 1214..1236 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" protein_bind 1251..1275 /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)." primer_bind complement(1275..1294) /label=T7 /note="T7 promoter, forward primer" promoter complement(1276..1294) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(1378..1397) /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" promoter 1603..1680 /gene="lacI" /label=lacI promoter CDS 1681..2763 /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 complement(1700..1719) /label=LacI-R /note="LacI, reverse primer" CDS 3572..3763 /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" primer_bind complement(3757..3779) /label=pGEX 3' /note="pGEX vectors, reverse primer" misc_feature 3865..4004 /label=bom /note="basis of mobility region from pBR322" primer_bind 3920..3939 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind complement(4019..4036) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4190..4778) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4270..4289) /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind complement(4889..4908) /label=pENTR-R /note="pENTR vectors, reverse primer" CDS 4900..5715 /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 complement(4973..4992) /label=Kan-R /note="Kanamycin resistance gene, reverse primer" rep_origin complement(5807..6262) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5944..5965) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 6156..6175 /label=F1ori-R /note="F1 origin, reverse primer" ORIGIN 1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa 61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt 121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagctttt 181 attaagcctc aggctcgcca gaaccttctg tacagccacg aattttttcc acgaaagcct 241 tagccttctc cgctacctcc acgggcgttc ccttaaccag tgccgatccg actcctacgg 301 caaggacacc ggctttaaac cactcacaga cattgtcaag atttacacca ccagtaggaa 361 cgaatttaac atttgggaat ggccctttca tagccttaac gaattgtggc cctacaactt 421 cccccggaaa caactttaat atagtgtgac ccagcttcat tgctttcact aactcggtag 481 gtgtcattac acctggcata taaaataccc ctttttcttt ggcaaattga ctgatctcct 541 cgtccaagtg cggggagact ataaattcag ctccgctttc cacagctttt ctggcttgct 601 caacggaagt gactgtccct gcaccaataa tggcccccat ttcctttaag aaggacaatt 661 ccttaataac agtatccgca tcaggtactg tgaacgtgat ctctataaga tgcacccctc 721 ccaagaacac tgctaaagcc tttttcttcg cttcctccac gctgtttgcg cgcaatactg 781 cgacaatctt atgtttcttg aaaagctctt ccattttcat tgaaccgcca gagccgccag 841 atccccctga gcccccagta tgagcgtcac cttcagttgc ttcaccatct acagtaacct 901 gaccgtcctc attaactgta aattcaattg gagttgctac ctcataaccg tctggtgctg 961 cggtttcgac aaatgtatat tttcctggat acaggtagaa atccttcaca tgtccatctg 1021 aaatccatgt actaatagtt ttaccagatg aatcacgcaa ctccatagtt gcaccagcta 1081 actcacggcc gtcctcatca cgttttgaga atttaatatg ggtagcacta tcttcttcag 1141 ttgtcatatc accggacgga ccttgttcac ctgataaacc tgataaggtg gttacacttg 1201 aacccatggt atatctcctt cttaaagtta aacaaaatta tttctagagg ggaattgtta 1261 tccgctcaca attcccctat agtgagtcgt attaatttcg cgggatcgag atctcgatcc 1321 tctacgccgg acgcatcgtg gccggcatca ccggcgccac aggtgcggtt gctggcgcct 1381 atatcgccga catcaccgat ggggaagatc gggctcgcca cttcgggctc atgagcgctt 1441 gtttcggcgt gggtatggtg gcaggccccg tggccggggg actgttgggc gccatctcct 1501 tgcatgcacc attccttgcg gcggcggtgc tcaacggcct caacctacta ctgggctgct 1561 tcctaatgca ggagtcgcat aagggagagc gtcgagatcc cggacaccat cgaatggcgc 1621 aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag ggtggtgaat 1681 gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt 1741 tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg 1801 gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc gggcaaacag 1861 tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc gcaaattgtc 1921 gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa 1981 cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt 2041 gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga agctgcctgc 2101 actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa cagtattatt 2161 ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt gggtcaccag 2221 caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg tctggctggc 2281 tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga aggcgactgg 2341 agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat cgttcccact 2401 gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc 2461 gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga agacagctca 2521 tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg gcaaaccagc 2581 gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca gctgttgccc 2641 gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc 2701 gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag 2761 tgagcgcaac gcaattaatg taagttagct cactcattag gcaccgggat ctcgaccgat 2821 gcccttgaga gccttcaacc cagtcagctc cttccggtgg gcgcggggca tgactatcgt 2881 cgccgcactt atgactgtct tctttatcat gcaactcgta ggacaggtgc cggcagcgct 2941 ctgggtcatt ttcggcgagg accgctttcg ctggagcgcg acgatgatcg gcctgtcgct 3001 tgcggtattc ggaatcttgc acgccctcgc tcaagccttc gtcactggtc ccgccaccaa 3061 acgtttcggc gagaagcagg ccattatcgc cggcatggcg gccccacggg tgcgcatgat 3121 cgtgctcctg tcgttgagga cccggctagg ctggcggggt tgccttactg gttagcagaa 3181 tgaatcaccg atacgcgagc gaacgtgaag cgactgctgc tgcaaaacgt ctgcgacctg 3241 agcaacaaca tgaatggtct tcggtttccg tgtttcgtaa agtctggaaa cgcggaagtc 3301 agcgccctgc accattatgt tccggatctg catcgcagga tgctgctggc taccctgtgg 3361 aacacctaca tctgtattaa cgaagcgctg gcattgaccc tgagtgattt ttctctggtc 3421 ccgccgcatc cataccgcca gttgtttacc ctcacaacgt tccagtaacc gggcatgttc 3481 atcatcagta acccgtatcg tgagcatcct ctctcgtttc atcggtatca ttacccccat 3541 gaacagaaat cccccttaca cggaggcatc agtgaccaaa caggaaaaaa ccgcccttaa 3601 catggcccgc tttatcagaa gccagacatt aacgcttctg gagaaactca acgagctgga 3661 cgcggatgaa caggcagaca tctgtgaatc gcttcacgac cacgctgatg agctttaccg 3721 cagctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga 3781 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc 3841 agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg atagcggagt 3901 gtatactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccattgcggt 3961 gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgctct tccgcttcct 4021 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa 4081 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa 4141 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc 4201 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga 4261 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc 4321 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt 4381 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct 4441 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg 4501 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta 4561 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct 4621 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa 4681 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt 4741 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta 4801 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgaacaata 4861 aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat tcaacgggaa 4921 acgtcttgct ctaggccgcg attaaattcc aacatggatg ctgatttata tgggtataaa 4981 tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct atcgattgta tgggaagccc 5041 gatgcgccag agttgtttct gaaacatggc aaaggtagcg ttgccaatga tgttacagat 5101 gagatggtca gactaaactg gctgacggaa tttatgcctc ttccgaccat caagcatttt 5161 atccgtactc ctgatgatgc atggttactc accactgcga tccccgggaa aacagcattc 5221 caggtattag aagaatatcc tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc 5281 ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt 5341 cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag tgattttgat 5401 gacgagcgta atggctggcc tgttgaacaa gtctggaaag aaatgcataa acttttgcca 5461 ttctcaccgg attcagtcgt cactcatggt gatttctcac ttgataacct tatttttgac 5521 gaggggaaat taataggttg tattgatgtt ggacgagtcg gaatcgcaga ccgataccag 5581 gatcttgcca tcctatggaa ctgcctcggt gagttttctc cttcattaca gaaacggctt 5641 tttcaaaaat atggtattga taatcctgat atgaataaat tgcagtttca tttgatgctc 5701 gatgagtttt tctaagaatt aattcatgag cggatacata tttgaatgta tttagaaaaa 5761 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt 5821 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag 5881 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt 5941 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga 6001 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg 6061 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct 6121 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc 6181 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt 6241 aatgcgccgc tacagggcgc gtcccattcg cca //