LOCUS Exported 6894 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expression construct for producing Avi tagged Tyrosyl-tRNA synthetase. ACCESSION . VERSION . KEYWORDS YARS_pNIC-CTB10H SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6894) TITLE Graslund lab plasmids REFERENCE 2 (bases 1 to 6894) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6894 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 971..990 /label=LacI-R /note="LacI, reverse primer" promoter complement(1010..1087) /gene="lacI" /label=lacI promoter primer_bind 1293..1312 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" primer_bind 1396..1415 /label=T7 /note="T7 promoter, forward primer" promoter 1396..1414 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 1415..1439 /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)." CDS 3070..3090 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 3097..3141 /codon_start=1 /product="peptide tag that allows for enzymatic biotinylation" /label=AviTag(TM) /translation="GLNDIFEAQKIEWHE" CDS 3142..3171 /codon_start=1 /product="10xHis affinity tag" /label=10xHis /translation="HHHHHHHHHH" primer_bind complement(3224..3242) /label=T7 Term /note="T7 terminator, reverse primer" terminator 3238..3285 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" rep_origin 3322..3777 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3409..3428) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3619..3640 /label=F1ori-F /note="F1 origin, forward primer" CDS complement(3869..4684) /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 4592..4611 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" primer_bind 4676..4695 /label=pENTR-R /note="pENTR vectors, reverse primer" rep_origin 4806..5394 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5295..5314 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5548..5565 /label=L4440 /note="L4440 vector, forward primer" misc_feature 5580..5719 /label=bom /note="basis of mobility region from pBR322" primer_bind complement(5645..5664) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 5805..5827 /label=pGEX 3' /note="pGEX vectors, reverse primer" CDS complement(5821..6012) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(6821..1009) /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" ORIGIN 1 cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct 61 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc 121 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt 181 cggtatcgtc gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa 241 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga 301 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt 361 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac 421 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca 481 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt 541 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt 601 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt 661 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg 721 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg 781 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg 841 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt 901 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat 961 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc 1021 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga 1081 tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt 1141 agtaggttga ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg 1201 cccaacagtc ccccggccac ggggcctgcc accataccca cgccgaaaca agcgctcatg 1261 agcccgaagt ggcgagcccg atcttcccca tcggtgatgt cggcgatata ggcgccagca 1321 accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag gatcgagatc 1381 tcgatcccgc gaaattaata cgactcacta taggggaatt gtgagcggat aacaattccc 1441 ctctagaaat aattttgttt aaccttaaga aggagatata ctatggggga cgctcccagc 1501 cctgaagaga aactgcacct tatcacccgg aacctgcagg aggttctggg ggaagagaag 1561 ctgaaggaga tactgaagga gcgggaactt aaaatttact ggggaacggc aaccacgggc 1621 aaaccacatg tggcttactt tgtgcccatg tcaaagattg cagacttctt aaaggcaggg 1681 tgtgaggtaa caattctgtt tgcggacctc cacgcatacc tggataacat gaaagcccca 1741 tgggaacttc tagaactccg agtcagttac tatgagaatg tgatcaaagc aatgctggag 1801 agcattggtg tgcccttgga gaagctcaag ttcatcaaag gcactgatta ccagctcagc 1861 aaagagtaca cactagatgt gtacagactc tcctccgtgg tcacacagca cgattccaag 1921 aaggctggag ctgaggtggt aaagcaggtg gagcaccctt tgctgagtgg cctcttatac 1981 cccggactgc aggctttgga tgaagagtat ttaaaagtag atgcccaatt tggaggcatt 2041 gatcagagaa agattttcac ctttgcagag aagtacctcc ctgcacttgg ctattcaaaa 2101 cgggtccatc tgatgaatcc tatggttcca ggattaacag gcagcaaaat gagctcttca 2161 gaagaggagt ccaagattga tctccttgat cggaaggagg atgtgaagaa aaaactgaag 2221 aaggccttct gtgagccagg aaatgtggag aacaatgggg ttctgtcctt catcaagcat 2281 gtcctttttc cccttaagtc cgagtttgtg atcctacgag atgagaaatg gggtggaaac 2341 aaaacctaca cagcttacgt ggacctggaa aaggactttg ctgctgaggt tgtacatcct 2401 ggagacctga agaattctgt tgaagtcgca ctgaacaagt tgctggatcc aatccgggaa 2461 aagtttaata cccctgccct gaaaaaactg gccagcgctg cctacccaga tccctcaaag 2521 cagaagccaa tggccaaagg ccctgccaag aattcagaac cagaggaggt catcccatcc 2581 cggctggata tccgtgtggg gaaaatcatc actgtggaga agcacccaga tgcagacagc 2641 ctgtatgtag agaagattga cgtgggggaa gctgaaccac ggactgtggt gagcggcctg 2701 gtacagttcg tgcccaagga ggaactgcag gacaggctgg tagtggtgct gtgcaacctg 2761 aaaccccaga agatgagagg agtcgagtcc caaggcatgc ttctgtgtgc ttctatagaa 2821 gggataaacc gccaggttga acctctggac cctccggcag gctctgctcc tggtgagcac 2881 gtgtttgtga agggctatga aaagggccaa ccagatgagg agctcaagcc caagaagaaa 2941 gtcttcgaga agttgcaggc tgacttcaaa atttctgagg agtgcatcgc acagtggaag 3001 caaaccaact tcatgaccaa gctgggctcc atttcctgta aatcgctgaa aggggggaac 3061 attagcgcag agaacctcta cttccaatcg ggtggcggtc tgaacgacat cttcgaggct 3121 cagaaaatcg aatggcacga acatcatcat catcaccacc accaccacca ctgagatccg 3181 gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga gcaataacta 3241 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa aggaggaact 3301 atatccggat tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg 3361 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt 3421 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc 3481 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg 3541 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg 3601 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct 3661 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg 3721 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg 3781 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc 3841 aaatatgtat ccgctcatga attaattctt agaaaaactc atcgagcatc aaatgaaact 3901 gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt ttctgtaatg 3961 aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat cggtctgcga 4021 ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa ataaggttat 4081 caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa agtttatgca 4141 tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa tcactcgcat 4201 caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg cgatcgctgt 4261 taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact gccagcgcat 4321 caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct gttttcccgg 4381 ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc ttgatggtcg 4441 gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta acatcattgg 4501 caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc ccatacaatc 4561 gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac ccatataaat 4621 cagcatccat gttggaattt aatcgcggcc tagagcaaga cgtttcccgt tgaatatggc 4681 tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt catgaccaaa 4741 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga 4801 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg 4861 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact 4921 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac 4981 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg 5041 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg 5101 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga 5161 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc 5221 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg 5281 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc 5341 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc 5401 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt 5461 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc 5521 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc 5581 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcaa tggtgcactc 5641 tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg 5701 tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc 5761 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg 5821 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc 5881 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt 5941 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc 6001 ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc 6061 gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact 6121 ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac 6181 tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca 6241 gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag 6301 actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt 6361 gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag 6421 gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg 6481 ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt 6541 gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat 6601 cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg 6661 tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg 6721 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg 6781 tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc 6841 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gagg //