LOCUS Exported 8310 bp ds-DNA circular SYN 09-JUN-2021 DEFINITION Expresses codon optimised full length His tagged PARP1 in bacterial cells. ACCESSION . VERSION . KEYWORDS PARP1 FL SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8310) AUTHORS Ogden TEH, Yang JC, Schimpl M, Easton LE, Underwood E, Rawlins PB, McCauley MM, Langelier MF, Pascal JM, Embrey KJ, Neuhaus D TITLE Dynamics of the HD regulatory subdomain of PARP-1; substrate access and allostery in PARP activation and inhibition. JOURNAL Nucleic Acids Res. 2021 Feb 26;49(4):2266-2288. doi: 10.1093/nar/gkab020. PUBMED 33511412 REFERENCE 2 (bases 1 to 8310) AUTHORS . TITLE Direct Submission JOURNAL Exported Jun 9, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8310 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 111..130 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" primer_bind 214..233 /label=T7 /note="T7 promoter, forward primer" promoter 214..232 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 233..257 /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 272..294 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" CDS 308..325 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS 3388..3405 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(3458..3476) /label=T7 Term /note="T7 terminator, reverse primer" terminator 3472..3519 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" rep_origin 3556..4011 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3643..3662) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3853..3874 /label=F1ori-F /note="F1 origin, forward primer" CDS complement(4103..4918) /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin in bacteria or G418 (Geneticin(R)) in eukaryotes" /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind 4826..4845 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" primer_bind 4910..4929 /label=pENTR-R /note="pENTR vectors, reverse primer" rep_origin 5040..5628 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5529..5548 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5782..5799 /label=L4440 /note="L4440 vector, forward primer" misc_feature 5814..5953 /label=bom /note="basis of mobility region from pBR322" primer_bind complement(5879..5898) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 6039..6061 /label=pGEX 3' /note="pGEX vectors, reverse primer" CDS complement(6055..6246) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(7055..8137) /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 8099..8118 /label=LacI-R /note="LacI, reverse primer" promoter complement(8138..8215) /gene="lacI" /label=lacI promoter ORIGIN 1 gcatgcaagg agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg 61 ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg 121 gcgatatagg cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg 181 gcgtagagga tcgagatctc gatcccgcga aattaatacg actcactata ggggaattgt 241 gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag gagatataca 301 tatgaaacac catcaccatc accatatgaa acaggcagaa agcagcgata aactgtatcg 361 tgtggaatat gcaaaaagcg gtcgtgcaag ctgtaaaaaa tgtagcgaga gcattccgaa 421 agatagcctg cgtatggcaa ttatggttca gagcccgatg tttgatggta aagtgccgca 481 ttggtatcat ttcagctgct tttggaaagt tggtcatagc attcgtcatc cggatgttga 541 agttgatggt tttagcgaac tgcgttggga tgatcagcag aaagtgaaaa aaaccgctga 601 agccggtggt gttactggta aaggtcagga tggtattggt agcaaagcag aaaaaaccct 661 gggtgatttt gcagcagaat atgccaaaag caatcgtagc acctgtaaag gctgcatgga 721 aaaaattgaa aaaggtcagg tgcgtctgag caaaaaaatg gttgatccgg aaaaaccgca 781 gctgggtatg attgatcgtt ggtatcatcc gggttgcttt gttaaaaatc gcgaagaact 841 gggttttcgt ccggaatata gcgcaagcca gctgaaaggt tttagcctgc tggcaaccga 901 agataaagaa gccctgaaaa aacagctgcc tggtgttaaa agcgaaggta aacgcaaagg 961 tgatgaagtg gatggtgttg atgaggttgc caaaaaaaaa agcaaaaaag agaaagataa 1021 agatagcaaa ctggaaaaag cactgaaagc ccagaatgat ctgatctgga acatcaaaga 1081 cgaactgaaa aaagtgtgta gcaccaacga tctgaaagaa ctgctgatct ttaataaaca 1141 gcaggttccg agcggtgaaa gcgcaattct ggatcgtgtt gccgatggta tggtttttgg 1201 tgcactgctg ccgtgtgaag aatgtagcgg tcagctggtt tttaaaagcg acgcatatta 1261 ttgtaccggt gatgttaccg catggaccaa atgtatggtt aaaacccaga ccccgaatcg 1321 taaagaatgg gtgaccccga aagaattccg tgagattagc tatctgaaaa aactgaaagt 1381 taaaaaacag gatcgcattt ttccgcctga aaccagcgca agcgttgcag caacaccgcc 1441 tccgagcacc gcatctgcac cggcagcagt taatagcagc gcaagcgcag ataaaccgct 1501 gagcaacatg aaaattctga ccctgggtaa actgagccgt aacaaagatg aagtgaaagc 1561 catgattgaa aaactgggtg gtaaactgac cggcaccgca aataaagcaa gcctgtgcat 1621 cagcaccaaa aaagaggtgg aaaaaatgaa taaaaaaatg gaagaagtga aagaagccaa 1681 tattcgcgtt gttagcgaag attttctgca agatgttagc gcaagcacca aaagcctgca 1741 agaactgttt ctggcacata ttctgagccc gtggggtgca gaagttaaag cagaaccggt 1801 tgaagttgtt gcaccgcgtg gtaaaagcgg tgcagcactg tctaaaaaat ccaaaggcca 1861 ggtgaaagaa gaaggcatca acaaaagcga aaaacgtatg aaactgaccc tgaaaggtgg 1921 tgcagcagtt gatccggata gcggtctgga acatagcgca catgttctgg aaaaaggtgg 1981 taaagttttt agcgcaaccc tgggtctggt tgatattgtt aaaggcacca atagctacta 2041 taaactgcaa ctgctggaag atgataaaga aaaccgctat tggatttttc gtagctgggg 2101 tcgtgttggc accgttattg gcagcaataa actggaacaa atgccgagca aagaagatgc 2161 catcgaacac ttcatgaaac tgtatgaaga aaaaaccggt aatgcctggc atagcaaaaa 2221 cttcaccaaa tatccgaaaa aattctatcc gctggaaatc gattatggtc aggatgaaga 2281 agcagttaaa aaactgaccg tgaatccggg tacaaaaagc aaactgccga aaccggttca 2341 ggatctgatc aaaatgatct ttgatgtgga aagcatgaaa aaagccatgg tggaatatga 2401 aattgacctg caaaaaatgc cgctgggcaa actgagcaaa cgtcagattc aggcagcata 2461 tagcattctg agcgaagttc agcaggcagt tagccagggt agcagcgata gccagattct 2521 ggatctgagc aatcgctttt ataccctgat tccgcacgat tttggtatga aaaaaccgcc 2581 tctgctgaat aatgcagata gcgttcaggc aaaagcagaa atgctggata acctgctgga 2641 tattgaagtt gcatatagcc tgctgcgtgg tggttctgat gatagcagca aagatccgat 2701 cgatgtgaac tatgagaaac tgaaaaccga tattaaagtg gttgatcgtg atagcgaaga 2761 agccgaaatt atccgcaaat acgtgaaaaa tacccatgca accacccata atgcctatga 2821 tctggaagtg attgatatct ttaaaatcga acgtgaaggt gaatgtcagc gctataaacc 2881 gtttaaacag ctgcataatc gtcgtctgct gtggcatggt agccgtacca ccaattttgc 2941 aggtattctg agccagggtc tgcgtattgc accgcctgaa gcaccggtta ccggttatat 3001 gtttggcaaa ggcatctatt ttgccgatat ggttagcaaa agcgccaatt attgtcatac 3061 cagccagggt gatccgattg gtctgattct gctgggtgaa gttgcactgg gtaacatgta 3121 tgaactgaaa catgccagcc atattagcaa actgcctaaa ggcaaacata gcgttaaagg 3181 tctgggcaaa accacaccgg atccgagcgc aaatattagc ctggatggtg tggatgttcc 3241 tctgggcacc ggtattagca gcggtgttaa tgataccagc ctgctgtata acgaatacat 3301 cgtgtatgat attgcccagg tgaatctgaa atacctgctg aaactgaaat tcaacttcaa 3361 aaccagcctg tggtagtaag actcgagcac caccaccacc accactgaga tccggctgct 3421 aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa 3481 ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc 3541 ggattggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta 3601 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc 3661 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt 3721 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg 3781 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca 3841 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct 3901 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga 3961 tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt taggtggcac 4021 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat 4081 gtatccgctc atgaattaat tcttagaaaa actcatcgag catcaaatga aactgcaatt 4141 tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt aatgaaggag 4201 aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct gcgattccga 4261 ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg ttatcaagtg 4321 agaaatcacc atgagtgacg actgaatccg gtgagaatgg caaaagttta tgcatttctt 4381 tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc gcatcaacca 4441 aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcgatcg ctgttaaaag 4501 gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa 4561 tattttcacc tgaatcagga tattcttcta atacctggaa tgctgttttc ccggggatcg 4621 cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg gtcggaagag 4681 gcataaattc cgtcagccag tttagtctga ccatctcatc tgtaacatca ttggcaacgc 4741 tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac aatcgataga 4801 ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat aaatcagcat 4861 ccatgttgga atttaatcgc ggcctagagc aagacgtttc ccgttgaata tggctcataa 4921 caccccttgt attactgttt atgtaagcag acagttttat tgttcatgac caaaatccct 4981 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct 5041 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca 5101 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc 5161 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc 5221 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct 5281 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag 5341 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc 5401 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg 5461 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag 5521 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt 5581 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac 5641 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg 5701 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc 5761 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg 5821 cggtattttc tccttacgca tctgtgcggt atttcacacc gcaatggtgc actctcagta 5881 caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg 5941 ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct 6001 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag 6061 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc 6121 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag 6181 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt 6241 ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa 6301 acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg 6361 ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg 6421 tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc 6481 tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta 6541 cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca 6601 gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc 6661 ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc 6721 catgccggcg ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa 6781 ggcttgagcg agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc 6841 gctccagcga aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac 6901 gagttgcatg ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca 6961 ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta 7021 atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa 7081 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat 7141 tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca 7201 ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa 7261 aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt 7321 atcccactac cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg 7381 cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca 7441 gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta 7501 tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg 7561 agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat 7621 gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct 7681 ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg 7741 catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat 7801 tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc 7861 tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca 7921 gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg 7981 ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt 8041 tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg 8101 catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct 8161 cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga 8221 tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg 8281 ccgttgagca ccgccgccgc aaggaatggt //