LOCUS Exported 6237 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Expresses Streptococcus pneumoniae PsaA (19-309) in bacterial cells. ACCESSION . VERSION . KEYWORDS PsaA SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6237) 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 6237) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6237 /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(1118..1138) /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQG" CDS complement(1148..1165) /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" RBS 1179..1201 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" protein_bind 1216..1240 /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(1240..1259) /label=T7 /note="T7 promoter, forward primer" promoter complement(1241..1259) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(1343..1362) /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" promoter 1568..1645 /gene="lacI" /label=lacI promoter CDS 1646..2728 /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(1665..1684) /label=LacI-R /note="LacI, reverse primer" CDS 3536..3727 /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" primer_bind complement(3721..3743) /label=pGEX 3' /note="pGEX vectors, reverse primer" misc_feature 3829..3968 /label=bom /note="basis of mobility region from pBR322" primer_bind 3884..3903 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind complement(3983..4000) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4154..4742) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4234..4253) /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind complement(4853..4872) /label=pENTR-R /note="pENTR vectors, reverse primer" CDS 4864..5679 /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(4937..4956) /label=Kan-R /note="Kanamycin resistance gene, reverse primer" rep_origin complement(5771..6226) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5908..5929) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 6120..6139 /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 ggtagtgctc gagtgcggcc gcaagctttt 181 attatttcgg cggtatcggt tcattggtga tataatgttt accgttttat tattttgcta 241 acccttctgc gattttgtcc agattatatt tcatcataga gtaataacta tcgccctctt 301 tcccttgctc ggcaatcgag tccgtaaaaa tttgcgcata gatagggata tttgtgtctt 361 gcgataccgt cttcatcggg cggtcatcca cagagctttc tacaaacagg ctaggaactt 421 ttgtctgacg aagtttttcg accagcgttt tgatttgttc cggggtgccc tcttcctcag 481 tgttgatctc ccaaatgtag gcggacggta cgccatatgc tttgctaaaa tatttgaacg 541 ctccctccga tgttacgatc aatttcttct cagccgggat cttgttgaac ttgtctttgg 601 attctttgtc caacttatcc aacttgtctg tgtactcttt cagatttttc tcatagaatt 661 ccttgttgtt cggatcttta gccgaaagtt gttttgcaat gtttttggca aagataatcc 721 cgttttcaag attcaaccag gcatgcgggt cctcttttcc tttttcgttt tgcccctcca 781 gatagattac gtctacacca tcactaacag caaaatagtc cttattctct gtttttttcg 841 cgttttctac caatttggtg aaccaagcat tgccaccggt ctccaaattg atcccattgt 901 agaagattaa atctgcctcg ctagtctttt tcacgtcttc tggaaggggt tcgtattcgt 961 gggggtcctg cccgatgggt acgattgaat gcaagtcaat tttgtcgccc gcaatgtttt 1021 tagtgatatc agcgataatt gaattcgtag ccacgacctt aagtttctgc cctgaagttg 1081 tatccttctt cccggatgcg caagcactac ccccactgcc ttggaaatac agattttctc 1141 cagacccatg atgatggtgg tgatgaccca tatgtatatc tccttcttaa agttaaacaa 1201 aattatttct agaggggaat tgttatccgc tcacaattcc cctatagtga gtcgtattaa 1261 tttcgcggga tcgagatctc gatcctctac gccggacgca tcgtggccgg catcaccggc 1321 gccacaggtg cggttgctgg cgcctatatc gccgacatca ccgatgggga agatcgggct 1381 cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg ccccgtggcc 1441 gggggactgt tgggcgccat ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac 1501 ggcctcaacc tactactggg ctgcttccta atgcaggagt cgcataaggg agagcgtcga 1561 gatcccggac accatcgaat ggcgcaaaac ctttcgcggt atggcatgat agcgcccgga 1621 agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg tcgcagagta 1681 tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc 1741 gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt 1801 ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc 1861 cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc 1921 cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca 1981 caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga 2041 tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga 2101 ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac tgggcgtgga 2161 gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt 2221 ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc 2281 gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat 2341 gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg 2401 cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg 2461 atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga 2521 ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc 2581 ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc 2641 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca 2701 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt tagctcactc 2761 attaggcacc gggatctcga ccgatgccct tgagagcctt caacccagtc agctccttcc 2821 ggtgggcgcg gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac 2881 tcgtaggaca ggtgccggca gcgctctggg tcatttcggc gaggaccgct ttcgctggag 2941 cgcgacgatg atcggcctgt cgcttgcggt attcggaatc ttgcacgccc tcgctcaagc 3001 cttcgtcact ggtcccgcca ccaaacgttt cggcgagaag caggccatta tcgccggcat 3061 ggcggcccca cgggtgcgca tgatcgtgct cctgtcgttg aggacccggc taggctggcg 3121 gggttgcctt actggttagc agaatgaatc accgatacgc gagcgaacgt gaagcgactg 3181 ctgctgcaaa acgtctgcga cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc 3241 gtaaagtctg gaaacgcgga agtcagcgcc ctgcaccatt atgttccgga tctgcatcgc 3301 aggatgctgc tggctaccct gtggaacacc tacatctgta ttaacgaagc gctggcattg 3361 accctgagtg atttttctct ggtcccgccg catccatacc gccagttgtt taccctcaca 3421 acgttccagt aaccgggcat gttcatcatc agtaacccgt atcgtgagca tcctctctcg 3481 tttcatcggt atcattaccc ccatgaacag aaatccccct tacacggagg catcagtgac 3541 caaacaggaa aaaaccgccc ttaacatggc ccgctttatc agaagccaga cattaacgct 3601 tctggagaaa ctcaacgagc tggacgcgga tgaacaggca gacatctgtg aatcgcttca 3661 cgaccacgct gatgagcttt accgcagctg cctcgcgcgt ttcggtgatg acggtgaaaa 3721 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag 3781 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac 3841 ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt 3901 gtactgagag tgcaccattg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc 3961 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc 4021 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata 4081 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg 4141 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct 4201 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa 4261 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc 4321 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt 4381 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg 4441 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg 4501 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct 4561 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc 4621 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg 4681 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc 4741 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt 4801 aagggatttt ggtcatgaac aataaaactg tctgcttaca taaacagtaa tacaaggggt 4861 gttatgagcc atattcaacg ggaaacgtct tgctctaggc cgcgattaaa ttccaacatg 4921 gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc aggtgcgaca 4981 atctatcgat tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca tggcaaaggt 5041 agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac ggaatttatg 5101 cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt actcaccact 5161 gcgatccccg ggaaaacagc attccaggta ttagaagaat atcctgattc aggtgaaaat 5221 attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt ttgtaattgt 5281 ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat gaataacggt 5341 ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga acaagtctgg 5401 aaagaaatgc ataaactttt gccattctca ccggattcag tcgtcactca tggtgatttc 5461 tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga tgttggacga 5521 gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct cggtgagttt 5581 tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc tgatatgaat 5641 aaattgcagt ttcatttgat gctcgatgag tttttctaag aattaattca tgagcggata 5701 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa 5761 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa 5821 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa 5881 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac 5941 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa 6001 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct 6061 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa 6121 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc 6181 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgcca //