LOCUS Exported 6708 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Expresses GST-RPN13 (19-132) R104A in E.coli. ACCESSION . VERSION . KEYWORDS pET151-GST-PP-RPN13 (19-132) R104A SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6708) AUTHORS Hemmis CW, Heard SC, Hill CP TITLE Phosphorylation of Tyr950 in the proteasome scaffolding protein RPN2 modulates its interaction with the ubiquitin receptor RPN13. JOURNAL J Biol Chem. 2019 May 7. pii: AC119.008881. doi: 10.1074/jbc.AC119.008881. PUBMED 31064842 REFERENCE 2 (bases 1 to 6708) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6708 /organism="synthetic DNA construct" /mol_type="other DNA" protein_bind 1..25 /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)." RBS 40..62 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" CDS 91..744 /codon_start=1 /product="glutathione S-transferase from Schistosoma japonicum" /label=GST /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK" primer_bind 702..724 /label=pGEX 5' /note="pGEX vectors, Glutathione-S-transferase, forward primer" CDS 751..774 /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /label=HRV 3C site /translation="LEVLFQGP" primer_bind complement(1182..1200) /label=T7 Term /note="T7 terminator, reverse primer" terminator 1196..1243 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" primer_bind complement(1481..1499) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 1567..1671 /gene="bla" /label=AmpR promoter CDS 1672..2532 /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 complement(1890..1909) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 2703..3291 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 3192..3211 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 3445..3462 /label=L4440 /note="L4440 vector, forward primer" misc_feature 3477..3616 /label=bom /note="basis of mobility region from pBR322" primer_bind complement(3542..3561) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 3702..3724 /label=pGEX 3' /note="pGEX vectors, reverse primer" CDS complement(3718..3909) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(5221..6303) /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 6265..6284 /label=LacI-R /note="LacI, reverse primer" promoter complement(6304..6381) /gene="lacI" /label=lacI promoter primer_bind 6587..6606 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" primer_bind 6690..1 /label=T7 /note="T7 promoter, forward primer" promoter 6690..6708 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" ORIGIN 1 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga 61 gatatacata tgggaattga tcccttcacc atgtccccta tactaggtta ttggaaaatt 121 aagggccttg tgcaacccac tcgacttctt ttggaatatc ttgaagaaaa atatgaagag 181 catttgtatg agcgcgatga aggtgataaa tggcgaaaca aaaagtttga attgggtttg 241 gagtttccca atcttcctta ttatattgat ggtgatgtta aattaacaca gtctatggcc 301 atcatacgtt atatagctga caagcacaac atgttgggtg gttgtccaaa agagcgtgca 361 gagatttcaa tgcttgaagg agcggttttg gatattagat acggtgtttc gagaattgca 421 tatagtaaag actttgaaac tctcaaagtt gattttctta gcaagctacc tgaaatgctg 481 aaaatgttcg aagatcgttt atgtcataaa acatatttaa atggtgatca tgtaacccat 541 cctgacttca tgttgtatga cgctcttgat gttgttttat acatggaccc aatgtgcctg 601 gatgcgttcc caaaattagt ttgttttaaa aaacgtattg aagctatccc acaaattgat 661 aagtacttga aatccagcaa gtatatagca tggcctttgc agggctggca agccacgttt 721 ggtggtggcg accatcctcc aaaatcggat ctggaagttc tgttccaggg gccgtccaac 781 aagtacttgg tggagtttcg ggcgggaaag atgtccctga aggggaccac cgtgactccg 841 gataagcgga aagggctggt gtacattcag cagacggacg actcgcttat tcacttctgc 901 tggaaggaca ggacgtccgg gaacgtggaa gacgacttga tcatcttccc tgacgactgt 961 gagttcaagc gggtgccgca gtgccccagc gggagggtct acgtgctgaa gttcaaggca 1021 gggtccaagg cgcttttctt ctggatgcag gaacccaaga cagaccagga tgaggagcat 1081 tgccggaaag tcaacgagta tctgaacaac cccccgtaaa agggcgagct cagatccggc 1141 tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc 1201 ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaag gaggaactat 1261 atccggatat cccgcaagag gcccggcagt accggcataa ccaagcctat gcctacagca 1321 tccagggtga cggtgccgag gatgacgatg agcgcattgt tagatttcat acacggtgcc 1381 tgactgcgtt agcaatttaa ctgtgataaa ctaccgcatt aaagctagct tatcgatgat 1441 aagctgtcaa acatgagaat taattcttga agacgaaagg gcctcgtgat acgcctattt 1501 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga 1561 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc 1621 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt 1681 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct 1741 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt 1801 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt 1861 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtgttgac 1921 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac 1981 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct 2041 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg 2101 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg 2161 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgcagca 2221 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa 2281 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt 2341 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc 2401 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg 2461 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt 2521 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt 2581 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc 2641 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct 2701 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta 2761 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc 2821 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac 2881 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct 2941 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat 3001 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg 3061 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa 3121 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg 3181 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga 3241 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc 3301 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct 3361 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct 3421 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg 3481 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcaatgg tgcactctca 3541 gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga 3601 ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg 3661 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca 3721 gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg 3781 gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc 3841 cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg ttttttcctg 3901 tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa tgataccgat 3961 gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc ggttactgga 4021 acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa aaatcactca 4081 gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta gccagcagca 4141 tcctgcgatg cagatccgga acataatggt gcagggcgct gacttccgcg tttccagact 4201 ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag acgttttgca 4261 gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac cagtaaggca 4321 accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca cccgtggcca 4381 ggacccaacg ctgcccgaga tgcgccgcgt gcggctgctg gagatggcgg acgcgatgga 4441 tatgttctgc caagggttgg tttgcgcatt cacagttctc cgcaagaatt gattggctcc 4501 aattcttgga gtggtgaatc cgttagcgag gtgccgccgg cttccattca ggtcgaggtg 4561 gcccggctcc atgcaccgcg acgcaacgcg gggaggcaga caaggtatag ggcggcgcct 4621 acaatccatg ccaacccgtt ccatgtgctc gccgaggcgg cataaatcgc cgtgacgatc 4681 agcggtccaa tgatcgaagt taggctggta agagccgcga gcgatccttg aagctgtccc 4741 tgatggtcgt catctacctg cctggacagc atggcctgca acgcgggcat cccgatgccg 4801 ccggaagcga gaagaatcat aatggggaag gccatccagc ctcgcgtcgc gaacgccagc 4861 aagacgtagc ccagcgcgtc ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa 4921 cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc 4981 gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc 5041 cagagcgctg ccggcacctg tcctacgagt tgcatgataa agaagacagt cataagtgcg 5101 gcgacgatag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag 5161 ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc 5221 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa 5281 cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga 5341 gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc 5401 cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata 5461 acatgagctg tcttcggtat cgtcgtatcc cactaccgag atatccgcac caacgcgcag 5521 cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat 5581 cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc 5641 actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg 5701 ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat 5761 ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga 5821 gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt 5881 agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag 5941 cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct 6001 tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc 6061 cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa 6121 cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat 6181 cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg 6241 ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt 6301 cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt 6361 tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac tcctgcatta 6421 ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat 6481 gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga 6541 aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga 6601 tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt 6661 agaggatcga gatctcgatc ccgcgaaatt aatacgactc actatagg //