LOCUS Exported 7464 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Protein expression and purification of ESRRB_Hormone-recep. ACCESSION . VERSION . KEYWORDS pHH0103_ESRRB_Hormone-recep SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7464) TITLE Sidhu Lab peptide-recognition modules (PRMs) REFERENCE 2 (bases 1 to 7464) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7464 /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 1659..1706 /label=fd terminator /note="central terminator from bacteriophage fd (Otsuka and Kunisawa, 1982)" rep_origin complement(3299..3754) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3436..3457) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3648..3667 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind complement(3796..3818) /label=pGEX 3' /note="pGEX vectors, reverse primer" misc_feature 3904..4044 /label=bom /note="basis of mobility region from pBR322" primer_bind 3959..3978 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind complement(4059..4076) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4230..4817) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(4988..5848) /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 5611..5630 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5849..5953) /gene="bla" /label=AmpR promoter primer_bind 6021..6039 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 6062..6139 /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 6140..7222 /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(6159..6178) /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 cggccatctc cccgccggct aaaaaaccgc tgacgaaaat tgtctcctac ctgctggtcg 961 cagaaccgga taaactgtat gctatgccgc cgccgggcat gccggaaggt gatatcaaag 1021 ccctgaccac gctgtgcgat ctggcagacc gtgaactggt ggttattatc ggctgggcta 1081 aacatattcc gggttttagc tctctgagcc tgggcgatca gatgtcactg ctgcaatcgg 1141 cctggatgga aattctgatc ctgggtattg tttatcgcag cctgccgtac gatgacaaac 1201 tggtctatgc ggaagattac atcatggacg aagaacactc tcgtctggct ggcctgctgg 1261 aactgtatcg cgcgattctg cagctggttc gtcgctacaa aaagctgaaa gtcgaaaagg 1321 aagaattcgt gaccctgaaa gcactggctc tggcgaacag tgattccatg tatatcgaag 1381 acctggaagc cgtgcagaag ctgcaagatc tgctgcatga agcactgcag gactacgaac 1441 tgtcccaacg tcacgaagaa ccgtggcgca ccggtaaact gctgctgacg ctgccgctgc 1501 tgcgtcagac ggcggccaag gcggttcaac atttttatag tgtcaaactg cagggcaagg 1561 tgccgatgca caaactgttc ctggaaatgc tggaagccaa ggcatgggct cgtgcggatt 1621 cactgcagga atggcgcccg ctggcggccg cataataaac cgatacaatt aaaggctcct 1681 tttggagcct tttttttgga gattttcaac gtgaaaaaat tattattcgc aattccttta 1741 gttgttcctt tctattctca ctccgctgaa actgttgaaa gttgtttagc aaaaccccat 1801 acagaaaatt catttactaa cgtctggaaa gacgacaaaa ctttagatcg ttacgctaac 1861 tatgagggct gtctgtggaa tgctacaggc gttgtagttt gtactggtga cgaaactcag 1921 tgtctagcta gagtggcggt ggctctggtt ccggtgattt tgattatgaa aagatggcaa 1981 acgctaataa gggggctatg accgaaaatg ccgatgaaaa cgcgctacag tctgacgcta 2041 aaggcaaact tgattctgtc gctactgatt acggtgctgc tatcgatggt ttcattggtg 2101 acgtttccgg ccttgctaat ggtaatggtg ctactggtga ttttgctggc tctaattccc 2161 aaatggctca agtcggtgac ggtgataatt cacctttaat gaataatttc cgtcaatatt 2221 taccttccct ccctcaatcg gttgaatgtc gcccttttgt ctttggcgct ggtaaaccat 2281 atgaattttc tattgattgt gacaaaataa acttattccg tggtgtcttt gcgtttcttt 2341 tatatgttgc cacctttatg tatgtatttt ctacgtttgc taacatactg cgtaataagg 2401 agtcttaatc atgccagttc ttttggctag cgccgcccta taccttgtct gcctccccgc 2461 gttgcgtcgc ggtgcatgga gccgggccac ctcgacctga atggaagccg gcggcacctc 2521 gctaacggat tcaccactcc aagaattgga gccaatcaat tcttgcggag aactgtgaat 2581 gcgcaaacca acccttggca gaacatatcc atcgcgtccg ccatctccag cagccgcacg 2641 cggcgcatct cgggcagcgt tgggtcctgg ccacgggtgc gcatgatcgt gctcctgtcg 2701 ttgaggaccc ggctaggctg gcggggttgc cttactggtt agcagaatga atcaccgata 2761 cgcgagcgaa cgtgaagcga ctgctgctgc aaaacgtctg cgacctgagc aacaacatga 2821 atggtcttcg gtttccgtgt ttcgtaaagt ctggaaacgc ggaagtcagc gccctgcacc 2881 attatgttcc ggatctgcat cgcaggatgc tgctggctac cctgtggaac acctacatct 2941 gtattaacga agcgctggca ttgaccctga gtgatttttc tctggtcccg ccgcatccat 3001 accgccagtt gtttaccctc acaacgttcc agtaaccggg catgttcatc atcagtaacc 3061 cgtatcgtga gcatcctctc tcgtttcatc ggtatcatta cccccatgaa cagaaatccc 3121 ccttacacgg aggcatcagt gaccaaacag gaaaaaaccg cccttaacat ggcccgcttt 3181 atcagaagcc agacattaac gcttctggag aaactcaacg agctggacgc ggatgaacag 3241 gcagacatct gtgaatcgct tcacgaccac gctgatgagc tttaccgcag gatccgcgaa 3301 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt 3361 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata 3421 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac 3481 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa 3541 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc 3601 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg 3661 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca 3721 cccgccgcgc ttaatgcgcc gctacagggc gcgtcgcgga tcctgcctcg cgcgtttcgg 3781 tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta 3841 agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg 3901 gggcgcagcc atgacccagt cacgtagcga tagcggagtg tatactggct taactatgcg 3961 gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc 4021 gtaaggagaa aataccgcat caggcgctct tccgcttcct cgctcactga ctcgctgcgc 4081 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc 4141 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg 4201 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat 4261 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag 4321 ggtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat 4381 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt 4441 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc 4501 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg 4561 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg 4621 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg 4681 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg 4741 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca 4801 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga 4861 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga 4921 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt 4981 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt 5041 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat 5101 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag 5161 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct 5221 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt 5281 tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg tttggtatgg 5341 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca 5401 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt 5461 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat 5521 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac 5581 cgagttgctc ttgcccggcg tcaacacggg ataataccgc gccacatagc agaactttaa 5641 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt 5701 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt 5761 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa 5821 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt 5881 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa 5941 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta 6001 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctt caagaattcc 6061 cgacaccatc gaatggtgca aaacctttcg cggtatggca tgatagcgcc cggaagagag 6121 tcaattcagg gtggtgaatg tgaaaccagt aacgttatac gatgtcgcag agtatgccgg 6181 tgtctcttat cagaccgttt cccgcgtggt gaaccaggcc agccacgttt ctgcgaaaac 6241 gcgggaaaaa gtggaagcgg cgatggcgga gctgaattac attcccaacc gcgtggcaca 6301 acaactggcg ggcaaacagt cgttgctgat tggcgttgcc acctccagtc tggccctgca 6361 cgcgccgtcg caaattgtcg cggcgattaa atctcgcgcc gatcaactgg gtgccagcgt 6421 ggtggtgtcg atggtagaac gaagcggcgt cgaagcctgt aaagcggcgg tgcacaatct 6481 tctcgcgcaa cgcgtcagtg ggctgatcat taactatccg ctggatgacc aggatgccat 6541 tgctgtggaa gctgcctgca ctaatgttcc ggcgttattt cttgatgtct ctgaccagac 6601 acccatcaac agtattattt tctcccatga agacggtacg cgactgggcg tggagcatct 6661 ggtcgcattg ggtcaccagc aaatcgcgct gttagcgggc ccattaagtt ctgtctcggc 6721 gcgtctgcgt ctggctggct ggcataaata tctcactcgc aatcaaattc agccgatagc 6781 ggaacgggaa ggcgactgga gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa 6841 tgagggcatc gttcccactg cgatgctggt tgccaacgat cagatggcgc tgggcgcaat 6901 gcgcgccatt accgagtccg ggctgcgcgt tggtgcggat atctcggtag tgggatacga 6961 cgataccgaa gacagctcat gttatatccc gccgttaacc accatcaaac aggattttcg 7021 cctgctgggg caaaccagcg tggaccgctt gctgcaactc tctcagggcc aggcggtgaa 7081 gggcaatcag ctgttgcccg tctcactggt gaaaagaaaa accaccctgg cgcccaatac 7141 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc 7201 ccgactggaa agcgggcagt gagcgcaacg caattaatgt aagttagctc actcattagg 7261 cacaattctc atgtttgaca gcttatcatc gactgcacgg tgcaccaatg cttctggcgt 7321 caggcagcca tcggaagctg tggtatggct gtgcaggtcg taaatcactg cataattcgt 7381 gtcgctcaag gcgcactccc gttctggata atgttttttg cgccgacatc ataacggttc 7441 tggcaaatat tctgaaatga gctg //