LOCUS Exported 7449 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Protein expression and purification of EEF1A2_GTP-EFTU. ACCESSION . VERSION . KEYWORDS pHH0103_EEF1A2_GTP-EFTU SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7449) TITLE Sidhu Lab peptide-recognition modules (PRMs) REFERENCE 2 (bases 1 to 7449) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7449 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 1..29 /label=tac promoter /note="strong E. coli promoter; hybrid between the trp and lac UV5 promoters" protein_bind 37..53 /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 42..64 /label=M13/pUC Reverse /note="In lacZ gene" CDS 138..155 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS 171..824 /codon_start=1 /product="glutathione S-transferase from Schistosoma japonicum" /label=GST /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK" primer_bind 782..804 /label=pGEX 5' /note="pGEX vectors, Glutathione-S-transferase, forward primer" CDS 831..854 /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /label=HRV 3C site /translation="LEVLFQGP" CDS 867..884 /codon_start=1 /product="thrombin recognition and cleavage site" /label=thrombin site /translation="LVPRGS" terminator 1644..1691 /label=fd terminator /note="central terminator from bacteriophage fd (Otsuka and Kunisawa, 1982)" rep_origin complement(3284..3739) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3421..3442) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3633..3652 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind complement(3781..3803) /label=pGEX 3' /note="pGEX vectors, reverse primer" misc_feature 3889..4029 /label=bom /note="basis of mobility region from pBR322" primer_bind 3944..3963 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind complement(4044..4061) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4215..4802) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(4973..5833) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 5596..5615 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5834..5938) /gene="bla" /label=AmpR promoter primer_bind 6006..6024 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 6047..6124 /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." CDS 6125..7207 /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(6144..6163) /label=LacI-R /note="LacI, reverse primer" ORIGIN 1 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca 61 caggaaacag ccagtccgtt taggtgtttt cacgagcact tcaccaacaa ggaccataga 121 ttatgaaaat cgaagaacac catcaccatc accattccag cggtaagctt atgtccccta 181 tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt ttggaatatc 241 ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa tggcgaaaca 301 aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat ggtgatgtta 361 aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac atgttgggtg 421 gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg gatattagat 481 acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt gattttctta 541 gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa acatatttaa 601 atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat gttgttttat 661 acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa aaacgtattg 721 aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca tggcctttgc 781 agggctggca agccacgttt ggtggtggcg accatcctcc aaaatcggat ctggaagttc 841 tgttccaggg gcccctgtcc agcggtctgg ttccgcgtgg ttccggtacc gcggcccagc 901 cggccggcaa agaaaagacc catattaaca tcgtggttat cggccacgtg gatagcggta 961 aatctaccac gaccggtcat ctgatttata aatgcggcgg tatcgacaag cgtaccattg 1021 aaaaatttga aaaggaagcg gccgaaatgg gcaaaggtag cttcaagtac gcgtgggttc 1081 tggataaact gaaggccgaa cgtgaacgcg gcattacgat cgacatttct ctgtggaaat 1141 ttgaaacgac caagtattac attaccatta tcgatgcacc gggccatcgc gacttcatca 1201 aaaatatgat tacgggtacc agtcaggcgg attgtgccgt tctgatcgtc gcagctggcg 1261 tgggtgaatt tgaagcgggc atttccaaaa acggtcagac gcgtgaacac gcactgctgg 1321 cttataccct gggcgttaaa caactgatcg tcggtgtgaa taaaatggat tcaacggaac 1381 cggcctattc ggaaaaacgc tacgacgaaa ttgtgaagga agttagcgcg tatatcaaaa 1441 agattggcta caacccggcc accgtcccgt ttgtgccgat cagcggctgg cacggtgata 1501 acatgctgga accgtctccg aatatgccgt ggttcaaagg ttggaaggtt gaacgtaaag 1561 aaggcaatgc aagtggtgtc tccctgctgg aagctctgga caccattctg ccgccgacgc 1621 gtccgaccgc ggccgcataa taaaccgata caattaaagg ctccttttgg agcctttttt 1681 ttggagattt tcaacgtgaa aaaattatta ttcgcaattc ctttagttgt tcctttctat 1741 tctcactccg ctgaaactgt tgaaagttgt ttagcaaaac cccatacaga aaattcattt 1801 actaacgtct ggaaagacga caaaacttta gatcgttacg ctaactatga gggctgtctg 1861 tggaatgcta caggcgttgt agtttgtact ggtgacgaaa ctcagtgtct agctagagtg 1921 gcggtggctc tggttccggt gattttgatt atgaaaagat ggcaaacgct aataaggggg 1981 ctatgaccga aaatgccgat gaaaacgcgc tacagtctga cgctaaaggc aaacttgatt 2041 ctgtcgctac tgattacggt gctgctatcg atggtttcat tggtgacgtt tccggccttg 2101 ctaatggtaa tggtgctact ggtgattttg ctggctctaa ttcccaaatg gctcaagtcg 2161 gtgacggtga taattcacct ttaatgaata atttccgtca atatttacct tccctccctc 2221 aatcggttga atgtcgccct tttgtctttg gcgctggtaa accatatgaa ttttctattg 2281 attgtgacaa aataaactta ttccgtggtg tctttgcgtt tcttttatat gttgccacct 2341 ttatgtatgt attttctacg tttgctaaca tactgcgtaa taaggagtct taatcatgcc 2401 agttcttttg gctagcgccg ccctatacct tgtctgcctc cccgcgttgc gtcgcggtgc 2461 atggagccgg gccacctcga cctgaatgga agccggcggc acctcgctaa cggattcacc 2521 actccaagaa ttggagccaa tcaattcttg cggagaactg tgaatgcgca aaccaaccct 2581 tggcagaaca tatccatcgc gtccgccatc tccagcagcc gcacgcggcg catctcgggc 2641 agcgttgggt cctggccacg ggtgcgcatg atcgtgctcc tgtcgttgag gacccggcta 2701 ggctggcggg gttgccttac tggttagcag aatgaatcac cgatacgcga gcgaacgtga 2761 agcgactgct gctgcaaaac gtctgcgacc tgagcaacaa catgaatggt cttcggtttc 2821 cgtgtttcgt aaagtctgga aacgcggaag tcagcgccct gcaccattat gttccggatc 2881 tgcatcgcag gatgctgctg gctaccctgt ggaacaccta catctgtatt aacgaagcgc 2941 tggcattgac cctgagtgat ttttctctgg tcccgccgca tccataccgc cagttgttta 3001 ccctcacaac gttccagtaa ccgggcatgt tcatcatcag taacccgtat cgtgagcatc 3061 ctctctcgtt tcatcggtat cattaccccc atgaacagaa atccccctta cacggaggca 3121 tcagtgacca aacaggaaaa aaccgccctt aacatggccc gctttatcag aagccagaca 3181 ttaacgcttc tggagaaact caacgagctg gacgcggatg aacaggcaga catctgtgaa 3241 tcgcttcacg accacgctga tgagctttac cgcaggatcc gcgaaattgt aagcgttaat 3301 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc 3361 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt 3421 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa 3481 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg 3541 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga 3601 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct 3661 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat 3721 gcgccgctac agggcgcgtc gcggatcctg cctcgcgcgt ttcggtgatg acggtgaaaa 3781 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag 3841 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac 3901 ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt 3961 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac 4021 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg 4081 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat 4141 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc 4201 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc 4261 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accagggttt ccccctggaa 4321 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc 4381 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt 4441 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg 4501 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg 4561 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct 4621 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc 4681 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg 4741 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc 4801 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt 4861 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa 4921 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat 4981 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct 5041 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg 5101 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag 5161 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta 5221 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg 5281 ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg 5341 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct 5401 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta 5461 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg 5521 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc 5581 cggcgtcaac acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg 5641 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga 5701 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg 5761 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat 5821 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc 5881 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca 5941 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct 6001 ataaaaatag gcgtatcacg aggccctttc gtcttcaaga attcccgaca ccatcgaatg 6061 gtgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt 6121 gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac 6181 cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga 6241 agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa 6301 acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat 6361 tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt 6421 agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt 6481 cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc 6541 ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat 6601 tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca 6661 ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc 6721 tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga 6781 ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc 6841 cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga 6901 gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag 6961 ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac 7021 cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt 7081 gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc 7141 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg 7201 gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcacaa ttctcatgtt 7261 tgacagctta tcatcgactg cacggtgcac caatgcttct ggcgtcaggc agccatcgga 7321 agctgtggta tggctgtgca ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca 7381 ctcccgttct ggataatgtt ttttgcgccg acatcataac ggttctggca aatattctga 7441 aatgagctg //