LOCUS Exported 7440 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Protein expression and purification of ESR2_Hormone-recep. ACCESSION . VERSION . KEYWORDS pHH0103_ESR2_Hormone-recep SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7440) TITLE Sidhu Lab peptide-recognition modules (PRMs) REFERENCE 2 (bases 1 to 7440) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7440 /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 1635..1682 /label=fd terminator /note="central terminator from bacteriophage fd (Otsuka and Kunisawa, 1982)" rep_origin complement(3275..3730) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3412..3433) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3624..3643 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind complement(3772..3794) /label=pGEX 3' /note="pGEX vectors, reverse primer" misc_feature 3880..4020 /label=bom /note="basis of mobility region from pBR322" primer_bind 3935..3954 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind complement(4035..4052) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4206..4793) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(4964..5824) /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 5587..5606 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5825..5929) /gene="bla" /label=AmpR promoter primer_bind 5997..6015 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 6038..6115 /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 6116..7198 /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(6135..6154) /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 cggccctgtc cccggaacaa ctggtgctga ccctgctgga agctgaaccg ccgcatgtcc 961 tgattagccg cccgtctgcc ccgtttaccg aagcgagcat gatgatgtct ctgaccaaac 1021 tggccgataa ggaactggtg catatgattt catgggcgaa aaagatcccg ggctttgttg 1081 aactgagcct gttcgaccaa gtccgtctgc tggaatcttg ctggatggaa gttctgatga 1141 tgggcctgat gtggcgctcc attgatcacc cgggtaaact gatctttgcc ccggacctgg 1201 tgctggatcg tgacgagggc aagtgcgttg aaggtattct ggaaatcttt gatatgctgc 1261 tggcaaccac gagccgtttc cgcgaactga aactgcaaca taaggaatat ctgtgtgtta 1321 aagctatgat tctgctgaac agctctatgt acccgctggt caccgcaacg caggatgcag 1381 acagttcccg caaactggcg cacctgctga atgcagtgac ggatgctctg gtctgggtga 1441 ttgccaaatc aggcatctca tcgcagcaac agtcgatgcg tctggcaaac ctgctgatgc 1501 tgctgagtca tgttcgccac gcttccaata agggtatgga acatctgctg aacatgaagt 1561 gtaagaatgt ggttccggtc tatgacctgc tgctggaaat gctgaacgcg cacgtgctgg 1621 cggccgcata ataaaccgat acaattaaag gctccttttg gagccttttt tttggagatt 1681 ttcaacgtga aaaaattatt attcgcaatt cctttagttg ttcctttcta ttctcactcc 1741 gctgaaactg ttgaaagttg tttagcaaaa ccccatacag aaaattcatt tactaacgtc 1801 tggaaagacg acaaaacttt agatcgttac gctaactatg agggctgtct gtggaatgct 1861 acaggcgttg tagtttgtac tggtgacgaa actcagtgtc tagctagagt ggcggtggct 1921 ctggttccgg tgattttgat tatgaaaaga tggcaaacgc taataagggg gctatgaccg 1981 aaaatgccga tgaaaacgcg ctacagtctg acgctaaagg caaacttgat tctgtcgcta 2041 ctgattacgg tgctgctatc gatggtttca ttggtgacgt ttccggcctt gctaatggta 2101 atggtgctac tggtgatttt gctggctcta attcccaaat ggctcaagtc ggtgacggtg 2161 ataattcacc tttaatgaat aatttccgtc aatatttacc ttccctccct caatcggttg 2221 aatgtcgccc ttttgtcttt ggcgctggta aaccatatga attttctatt gattgtgaca 2281 aaataaactt attccgtggt gtctttgcgt ttcttttata tgttgccacc tttatgtatg 2341 tattttctac gtttgctaac atactgcgta ataaggagtc ttaatcatgc cagttctttt 2401 ggctagcgcc gccctatacc ttgtctgcct ccccgcgttg cgtcgcggtg catggagccg 2461 ggccacctcg acctgaatgg aagccggcgg cacctcgcta acggattcac cactccaaga 2521 attggagcca atcaattctt gcggagaact gtgaatgcgc aaaccaaccc ttggcagaac 2581 atatccatcg cgtccgccat ctccagcagc cgcacgcggc gcatctcggg cagcgttggg 2641 tcctggccac gggtgcgcat gatcgtgctc ctgtcgttga ggacccggct aggctggcgg 2701 ggttgcctta ctggttagca gaatgaatca ccgatacgcg agcgaacgtg aagcgactgc 2761 tgctgcaaaa cgtctgcgac ctgagcaaca acatgaatgg tcttcggttt ccgtgtttcg 2821 taaagtctgg aaacgcggaa gtcagcgccc tgcaccatta tgttccggat ctgcatcgca 2881 ggatgctgct ggctaccctg tggaacacct acatctgtat taacgaagcg ctggcattga 2941 ccctgagtga tttttctctg gtcccgccgc atccataccg ccagttgttt accctcacaa 3001 cgttccagta accgggcatg ttcatcatca gtaacccgta tcgtgagcat cctctctcgt 3061 ttcatcggta tcattacccc catgaacaga aatccccctt acacggaggc atcagtgacc 3121 aaacaggaaa aaaccgccct taacatggcc cgctttatca gaagccagac attaacgctt 3181 ctggagaaac tcaacgagct ggacgcggat gaacaggcag acatctgtga atcgcttcac 3241 gaccacgctg atgagcttta ccgcaggatc cgcgaaattg taagcgttaa tattttgtta 3301 aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc 3361 aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg 3421 aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat 3481 cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc 3541 cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag 3601 ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg 3661 gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta 3721 cagggcgcgt cgcggatcct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca 3781 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc 3841 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg 3901 tagcgatagc ggagtgtata ctggcttaac tatgcggcat cagagcagat tgtactgaga 3961 gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg 4021 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg 4081 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga 4141 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg 4201 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag 4261 aggtggcgaa acccgacagg actataaaga taccagggtt tccccctgga agctccctcg 4321 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg 4381 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc 4441 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg 4501 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca 4561 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt 4621 ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag 4681 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg 4741 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc 4801 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt 4861 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt 4921 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca 4981 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg 5041 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac 5101 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg 5161 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc 5221 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgctg 5281 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac 5341 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc 5401 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac 5461 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact 5521 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa 5581 cacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt 5641 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca 5701 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa 5761 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac 5821 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg 5881 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc 5941 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata 6001 ggcgtatcac gaggcccttt cgtcttcaag aattcccgac accatcgaat ggtgcaaaac 6061 ctttcgcggt atggcatgat agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa 6121 accagtaacg ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg 6181 cgtggtgaac caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat 6241 ggcggagctg aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt 6301 gctgattggc gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc 6361 gattaaatct cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag 6421 cggcgtcgaa gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct 6481 gatcattaac tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa 6541 tgttccggcg ttatttcttg atgtctctga ccagacaccc atcaacagta ttattttctc 6601 ccatgaagac ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc accagcaaat 6661 cgcgctgtta gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca 6721 taaatatctc actcgcaatc aaattcagcc gatagcggaa cgggaaggcg actggagtgc 6781 catgtccggt tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat 6841 gctggttgcc aacgatcaga tggcgctggg cgcaatgcgc gccattaccg agtccgggct 6901 gcgcgttggt gcggatatct cggtagtggg atacgacgat accgaagaca gctcatgtta 6961 tatcccgccg ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga 7021 ccgcttgctg caactctctc agggccaggc ggtgaagggc aatcagctgt tgcccgtctc 7081 actggtgaaa agaaaaacca ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt 7141 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc 7201 gcaacgcaat taatgtaagt tagctcactc attaggcaca attctcatgt ttgacagctt 7261 atcatcgact gcacggtgca ccaatgcttc tggcgtcagg cagccatcgg aagctgtggt 7321 atggctgtgc aggtcgtaaa tcactgcata attcgtgtcg ctcaaggcgc actcccgttc 7381 tggataatgt tttttgcgcc gacatcataa cggttctggc aaatattctg aaatgagctg //