LOCUS Exported 6969 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Protein expression and purification of DLG4_PDZ_3. ACCESSION . VERSION . KEYWORDS pHH0103_DLG4_PDZ_3 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6969) TITLE Sidhu Lab peptide-recognition modules (PRMs) REFERENCE 2 (bases 1 to 6969) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6969 /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 1164..1211 /label=fd terminator /note="central terminator from bacteriophage fd (Otsuka and Kunisawa, 1982)" rep_origin complement(2804..3259) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2941..2962) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3153..3172 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind complement(3301..3323) /label=pGEX 3' /note="pGEX vectors, reverse primer" misc_feature 3409..3549 /label=bom /note="basis of mobility region from pBR322" primer_bind 3464..3483 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind complement(3564..3581) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(3735..4322) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(4493..5353) /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 5116..5135 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5354..5458) /gene="bla" /label=AmpR promoter primer_bind 5526..5544 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 5567..5644 /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 5645..6727 /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(5664..5683) /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 cggcccgtat tgtcatccat cgcggcagca ccggcctggg ttttaacatt gtgggcggtg 961 aagacggcga aggtattttt atcagtttca tcctggcagg cggtccggct gatctgtccg 1021 gcgaactgcg taaaggtgac cagattctga gcgtgaacgg cgttgatctg cgcaatgcat 1081 ctcacgaaca agcggccatc gcgctgaaaa atgccggtca gaccgttacg attatcgccc 1141 aatataaagc ggccgcataa taaaccgata caattaaagg ctccttttgg agcctttttt 1201 ttggagattt tcaacgtgaa aaaattatta ttcgcaattc ctttagttgt tcctttctat 1261 tctcactccg ctgaaactgt tgaaagttgt ttagcaaaac cccatacaga aaattcattt 1321 actaacgtct ggaaagacga caaaacttta gatcgttacg ctaactatga gggctgtctg 1381 tggaatgcta caggcgttgt agtttgtact ggtgacgaaa ctcagtgtct agctagagtg 1441 gcggtggctc tggttccggt gattttgatt atgaaaagat ggcaaacgct aataaggggg 1501 ctatgaccga aaatgccgat gaaaacgcgc tacagtctga cgctaaaggc aaacttgatt 1561 ctgtcgctac tgattacggt gctgctatcg atggtttcat tggtgacgtt tccggccttg 1621 ctaatggtaa tggtgctact ggtgattttg ctggctctaa ttcccaaatg gctcaagtcg 1681 gtgacggtga taattcacct ttaatgaata atttccgtca atatttacct tccctccctc 1741 aatcggttga atgtcgccct tttgtctttg gcgctggtaa accatatgaa ttttctattg 1801 attgtgacaa aataaactta ttccgtggtg tctttgcgtt tcttttatat gttgccacct 1861 ttatgtatgt attttctacg tttgctaaca tactgcgtaa taaggagtct taatcatgcc 1921 agttcttttg gctagcgccg ccctatacct tgtctgcctc cccgcgttgc gtcgcggtgc 1981 atggagccgg gccacctcga cctgaatgga agccggcggc acctcgctaa cggattcacc 2041 actccaagaa ttggagccaa tcaattcttg cggagaactg tgaatgcgca aaccaaccct 2101 tggcagaaca tatccatcgc gtccgccatc tccagcagcc gcacgcggcg catctcgggc 2161 agcgttgggt cctggccacg ggtgcgcatg atcgtgctcc tgtcgttgag gacccggcta 2221 ggctggcggg gttgccttac tggttagcag aatgaatcac cgatacgcga gcgaacgtga 2281 agcgactgct gctgcaaaac gtctgcgacc tgagcaacaa catgaatggt cttcggtttc 2341 cgtgtttcgt aaagtctgga aacgcggaag tcagcgccct gcaccattat gttccggatc 2401 tgcatcgcag gatgctgctg gctaccctgt ggaacaccta catctgtatt aacgaagcgc 2461 tggcattgac cctgagtgat ttttctctgg tcccgccgca tccataccgc cagttgttta 2521 ccctcacaac gttccagtaa ccgggcatgt tcatcatcag taacccgtat cgtgagcatc 2581 ctctctcgtt tcatcggtat cattaccccc atgaacagaa atccccctta cacggaggca 2641 tcagtgacca aacaggaaaa aaccgccctt aacatggccc gctttatcag aagccagaca 2701 ttaacgcttc tggagaaact caacgagctg gacgcggatg aacaggcaga catctgtgaa 2761 tcgcttcacg accacgctga tgagctttac cgcaggatcc gcgaaattgt aagcgttaat 2821 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc 2881 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt 2941 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa 3001 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg 3061 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga 3121 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct 3181 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat 3241 gcgccgctac agggcgcgtc gcggatcctg cctcgcgcgt ttcggtgatg acggtgaaaa 3301 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag 3361 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac 3421 ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt 3481 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac 3541 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg 3601 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat 3661 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc 3721 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc 3781 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accagggttt ccccctggaa 3841 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc 3901 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt 3961 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg 4021 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg 4081 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct 4141 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc 4201 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg 4261 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc 4321 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt 4381 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa 4441 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat 4501 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct 4561 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg 4621 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag 4681 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta 4741 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg 4801 ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg 4861 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct 4921 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta 4981 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg 5041 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc 5101 cggcgtcaac acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg 5161 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga 5221 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg 5281 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat 5341 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc 5401 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca 5461 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct 5521 ataaaaatag gcgtatcacg aggccctttc gtcttcaaga attcccgaca ccatcgaatg 5581 gtgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt 5641 gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac 5701 cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga 5761 agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa 5821 acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat 5881 tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt 5941 agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt 6001 cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc 6061 ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat 6121 tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca 6181 ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc 6241 tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga 6301 ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc 6361 cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga 6421 gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag 6481 ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac 6541 cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt 6601 gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc 6661 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg 6721 gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcacaa ttctcatgtt 6781 tgacagctta tcatcgactg cacggtgcac caatgcttct ggcgtcaggc agccatcgga 6841 agctgtggta tggctgtgca ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca 6901 ctcccgttct ggataatgtt ttttgcgccg acatcataac ggttctggca aatattctga 6961 aatgagctg //