LOCUS Exported 6987 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Protein expression and purification of DLG4_PDZ_2. ACCESSION . VERSION . KEYWORDS pHH0103_DLG4_PDZ_2 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6987) TITLE Sidhu Lab peptide-recognition modules (PRMs) REFERENCE 2 (bases 1 to 6987) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6987 /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 1182..1229 /label=fd terminator /note="central terminator from bacteriophage fd (Otsuka and Kunisawa, 1982)" rep_origin complement(2822..3277) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2959..2980) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3171..3190 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind complement(3319..3341) /label=pGEX 3' /note="pGEX vectors, reverse primer" misc_feature 3427..3567 /label=bom /note="basis of mobility region from pBR322" primer_bind 3482..3501 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind complement(3582..3599) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(3753..4340) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(4511..5371) /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 5134..5153 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5372..5476) /gene="bla" /label=AmpR promoter primer_bind 5544..5562 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 5585..5662 /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 5663..6745 /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(5682..5701) /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 cggccgaaat taaactgatc aaaggtccga aaggcctggg ttttagcatt gcaggcggtg 961 tgggcaacca gcatattccg ggtgataact ctatctacgt taccaaaatc atcgaaggcg 1021 gtgcggccca caaagatggc cgtctgcaaa ttggtgacaa aatcctggcg gtcaacagtg 1081 tgggtctgga agatgttatg catgaagacg ccgtcgcagc tctgaaaaat acgtatgacg 1141 tggtttacct gaaagtcgcc aaaccggcgg ccgcataata aaccgataca attaaaggct 1201 ccttttggag cctttttttt ggagattttc aacgtgaaaa aattattatt cgcaattcct 1261 ttagttgttc ctttctattc tcactccgct gaaactgttg aaagttgttt agcaaaaccc 1321 catacagaaa attcatttac taacgtctgg aaagacgaca aaactttaga tcgttacgct 1381 aactatgagg gctgtctgtg gaatgctaca ggcgttgtag tttgtactgg tgacgaaact 1441 cagtgtctag ctagagtggc ggtggctctg gttccggtga ttttgattat gaaaagatgg 1501 caaacgctaa taagggggct atgaccgaaa atgccgatga aaacgcgcta cagtctgacg 1561 ctaaaggcaa acttgattct gtcgctactg attacggtgc tgctatcgat ggtttcattg 1621 gtgacgtttc cggccttgct aatggtaatg gtgctactgg tgattttgct ggctctaatt 1681 cccaaatggc tcaagtcggt gacggtgata attcaccttt aatgaataat ttccgtcaat 1741 atttaccttc cctccctcaa tcggttgaat gtcgcccttt tgtctttggc gctggtaaac 1801 catatgaatt ttctattgat tgtgacaaaa taaacttatt ccgtggtgtc tttgcgtttc 1861 ttttatatgt tgccaccttt atgtatgtat tttctacgtt tgctaacata ctgcgtaata 1921 aggagtctta atcatgccag ttcttttggc tagcgccgcc ctataccttg tctgcctccc 1981 cgcgttgcgt cgcggtgcat ggagccgggc cacctcgacc tgaatggaag ccggcggcac 2041 ctcgctaacg gattcaccac tccaagaatt ggagccaatc aattcttgcg gagaactgtg 2101 aatgcgcaaa ccaacccttg gcagaacata tccatcgcgt ccgccatctc cagcagccgc 2161 acgcggcgca tctcgggcag cgttgggtcc tggccacggg tgcgcatgat cgtgctcctg 2221 tcgttgagga cccggctagg ctggcggggt tgccttactg gttagcagaa tgaatcaccg 2281 atacgcgagc gaacgtgaag cgactgctgc tgcaaaacgt ctgcgacctg agcaacaaca 2341 tgaatggtct tcggtttccg tgtttcgtaa agtctggaaa cgcggaagtc agcgccctgc 2401 accattatgt tccggatctg catcgcagga tgctgctggc taccctgtgg aacacctaca 2461 tctgtattaa cgaagcgctg gcattgaccc tgagtgattt ttctctggtc ccgccgcatc 2521 cataccgcca gttgtttacc ctcacaacgt tccagtaacc gggcatgttc atcatcagta 2581 acccgtatcg tgagcatcct ctctcgtttc atcggtatca ttacccccat gaacagaaat 2641 cccccttaca cggaggcatc agtgaccaaa caggaaaaaa ccgcccttaa catggcccgc 2701 tttatcagaa gccagacatt aacgcttctg gagaaactca acgagctgga cgcggatgaa 2761 caggcagaca tctgtgaatc gcttcacgac cacgctgatg agctttaccg caggatccgc 2821 gaaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca 2881 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag 2941 atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc 3001 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc 3061 taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc 3121 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa 3181 gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc 3241 acacccgccg cgcttaatgc gccgctacag ggcgcgtcgc ggatcctgcc tcgcgcgttt 3301 cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct 3361 gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg 3421 tcggggcgca gccatgaccc agtcacgtag cgatagcgga gtgtatactg gcttaactat 3481 gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat accgcacaga 3541 tgcgtaagga gaaaataccg catcaggcgc tcttccgctt cctcgctcac tgactcgctg 3601 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta 3661 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc 3721 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag 3781 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac 3841 cagggtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg 3901 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta 3961 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg 4021 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac 4081 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag 4141 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat 4201 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat 4261 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc 4321 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt 4381 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct 4441 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt 4501 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc 4561 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac 4621 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat 4681 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg 4741 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata 4801 gtttgcgcaa cgttgttgcc attgctgcag gcatcgtggt gtcacgctcg tcgtttggta 4861 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt 4921 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag 4981 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa 5041 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc 5101 gaccgagttg ctcttgcccg gcgtcaacac gggataatac cgcgccacat agcagaactt 5161 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc 5221 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta 5281 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa 5341 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca 5401 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac 5461 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta 5521 ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt cttcaagaat 5581 tcccgacacc atcgaatggt gcaaaacctt tcgcggtatg gcatgatagc gcccggaaga 5641 gagtcaattc agggtggtga atgtgaaacc agtaacgtta tacgatgtcg cagagtatgc 5701 cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag gccagccacg tttctgcgaa 5761 aacgcgggaa aaagtggaag cggcgatggc ggagctgaat tacattccca accgcgtggc 5821 acaacaactg gcgggcaaac agtcgttgct gattggcgtt gccacctcca gtctggccct 5881 gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac tgggtgccag 5941 cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg cggtgcacaa 6001 tcttctcgcg caacgcgtca gtgggctgat cattaactat ccgctggatg accaggatgc 6061 cattgctgtg gaagctgcct gcactaatgt tccggcgtta tttcttgatg tctctgacca 6121 gacacccatc aacagtatta ttttctccca tgaagacggt acgcgactgg gcgtggagca 6181 tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa gttctgtctc 6241 ggcgcgtctg cgtctggctg gctggcataa atatctcact cgcaatcaaa ttcagccgat 6301 agcggaacgg gaaggcgact ggagtgccat gtccggtttt caacaaacca tgcaaatgct 6361 gaatgagggc atcgttccca ctgcgatgct ggttgccaac gatcagatgg cgctgggcgc 6421 aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg gatatctcgg tagtgggata 6481 cgacgatacc gaagacagct catgttatat cccgccgtta accaccatca aacaggattt 6541 tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg gccaggcggt 6601 gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga aaaaccaccc tggcgcccaa 6661 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt 6721 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtaagttag ctcactcatt 6781 aggcacaatt ctcatgtttg acagcttatc atcgactgca cggtgcacca atgcttctgg 6841 cgtcaggcag ccatcggaag ctgtggtatg gctgtgcagg tcgtaaatca ctgcataatt 6901 cgtgtcgctc aaggcgcact cccgttctgg ataatgtttt ttgcgccgac atcataacgg 6961 ttctggcaaa tattctgaaa tgagctg //