LOCUS       Exported                7857 bp ds-DNA     circular SYN 30-SEP-2021
DEFINITION  Bacterial expression of PARP1 lacking Zn3 domain but containing 
            L713F gain-of-function mutation.
ACCESSION   .
VERSION     .
KEYWORDS    pET28_6xHis-PARP1delZn3-L713F
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7857)
  AUTHORS   Ooi SK, Sato S, Tomomori-Sato C, Zhang Y, Wen Z, Banks CAS, Washburn
            MP, Unruh JR, Florens L, Conaway RC, Conaway JW
  TITLE     Multiple roles for PARP1 in ALC1-dependent nucleosome remodeling.
  JOURNAL   Proc Natl Acad Sci U S A. 2021 Sep 7;118(36). pii: 2107277118. doi: 
            10.1073/pnas.2107277118.
  PUBMED    34465625
REFERENCE   2  (bases 1 to 7857)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 30, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7857
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             1426..1617
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1611..1633)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    1719..1858
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     1774..1793
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1873..1890)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2044..2632)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2124..2143)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(2743..2762)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2754..3569
                     /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     complement(2827..2846)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(3661..4116)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3798..3819)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4010..4029
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      4153..4200
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     4196..4214
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(4267..4284)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(6862..6879)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(6889..6906)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             6925..6947
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    6962..6986
                     /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     complement(6986..7005)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6987..7005)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7089..7108)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        7314..7391
                     /gene="lacI"
                     /label=lacI promoter
     CDS             7392..617
                     /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(7411..7430)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 tcaacagtat tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg
       61 cattgggtca ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc
      121 tgcgtctggc tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac
      181 gggaaggcga ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg
      241 gcatcgttcc cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg
      301 ccattaccga gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata
      361 ccgaagacag ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc
      421 tggggcaaac cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca
      481 atcagctgtt gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa
      541 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
      601 tggaaagcgg gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcaccg
      661 ggatctcgac cgatgccctt gagagccttc aacccagtca gctccttccg gtgggcgcgg
      721 ggcatgacta tcgtcgccgc acttatgact gtcttcttta tcatgcaact cgtaggacag
      781 gtgccggcag cgctctgggt cattttcggc gaggaccgct ttcgctggag cgcgacgatg
      841 atcggcctgt cgcttgcggt attcggaatc ttgcacgccc tcgctcaagc cttcgtcact
      901 ggtcccgcca ccaaacgttt cggcgagaag caggccatta tcgccggcat ggcggcccca
      961 cgggtgcgca tgatcgtgct cctgtcgttg aggacccggc taggctggcg gggttgcctt
     1021 actggttagc agaatgaatc accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa
     1081 acgtctgcga cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg
     1141 gaaacgcgga agtcagcgcc ctgcaccatt atgttccgga tctgcatcgc aggatgctgc
     1201 tggctaccct gtggaacacc tacatctgta ttaacgaagc gctggcattg accctgagtg
     1261 atttttctct ggtcccgccg catccatacc gccagttgtt taccctcaca acgttccagt
     1321 aaccgggcat gttcatcatc agtaacccgt atcgtgagca tcctctctcg tttcatcggt
     1381 atcattaccc ccatgaacag aaatccccct tacacggagg catcagtgac caaacaggaa
     1441 aaaaccgccc ttaacatggc ccgctttatc agaagccaga cattaacgct tctggagaaa
     1501 ctcaacgagc tggacgcgga tgaacaggca gacatctgtg aatcgcttca cgaccacgct
     1561 gatgagcttt accgcagctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     1621 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     1681 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt
     1741 agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt gtactgagag
     1801 tgcaccattg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg
     1861 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     1921 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     1981 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     2041 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     2101 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     2161 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     2221 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     2281 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     2341 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     2401 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     2461 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt
     2521 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     2581 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     2641 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     2701 ggtcatgaac aataaaactg tctgcttaca taaacagtaa tacaaggggt gttatgagcc
     2761 atattcaacg ggaaacgtct tgctctaggc cgcgattaaa ttccaacatg gatgctgatt
     2821 tatatgggta taaatgggct cgcgataatg tcgggcaatc aggtgcgaca atctatcgat
     2881 tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca
     2941 atgatgttac agatgagatg gtcagactaa actggctgac ggaatttatg cctcttccga
     3001 ccatcaagca ttttatccgt actcctgatg atgcatggtt actcaccact gcgatccccg
     3061 ggaaaacagc attccaggta ttagaagaat atcctgattc aggtgaaaat attgttgatg
     3121 cgctggcagt gttcctgcgc cggttgcatt cgattcctgt ttgtaattgt ccttttaaca
     3181 gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat gaataacggt ttggttgatg
     3241 cgagtgattt tgatgacgag cgtaatggct ggcctgttga acaagtctgg aaagaaatgc
     3301 ataaactttt gccattctca ccggattcag tcgtcactca tggtgatttc tcacttgata
     3361 accttatttt tgacgagggg aaattaatag gttgtattga tgttggacga gtcggaatcg
     3421 cagaccgata ccaggatctt gccatcctat ggaactgcct cggtgagttt tctccttcat
     3481 tacagaaacg gctttttcaa aaatatggta ttgataatcc tgatatgaat aaattgcagt
     3541 ttcatttgat gctcgatgag tttttctaag aattaattca tgagcggata catatttgaa
     3601 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     3661 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     3721 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     3781 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     3841 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     3901 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     3961 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     4021 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     4081 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccaatc cggatatagt
     4141 tcctcctttc agcaaaaaac ccctcaagac ccgtttagag gccccaaggg gttatgctag
     4201 ttattgctca gcggtggcag cagccaactc agcttccttt cgggctttgt tagcagccgg
     4261 atctcagtgg tggtggtggt ggtgctcgag ttaccacagg gaggtcttaa aattgaattt
     4321 cagtttcagc agatacttca gatttacctg agcaatatca tagacaatgt actcgttata
     4381 tagtagagag gtgtcattca caccagatga aatcccggtc ccaagaggaa cgtctacacc
     4441 atccagacta atgttagctg aaggatcagg ggtagttttg cccaaacctt tgacactgtg
     4501 cttgcccttg ggtaacttgc tgatatgtga agcgtgcttc agttcataca tgtttccaag
     4561 ggcaacttct cccaacagga ttaagcctat tgggtctccc tgagacgtat ggcagtagtt
     4621 ggcactcttg gagaccatgt cagcgaaata gatcccttta ccaaacatgt agcctgtcac
     4681 gggcgcttca ggcggggcta tccgaagacc ctgggacagg atcccagcaa agttggtggt
     4741 cctggacccg tgccacagca atcttcggtt atgaagctgc ttaaagggct tgtaacgctg
     4801 gcattcgcct tcacgctcta tcttaaagat atcgatgact tccaagtcat acgcattgtg
     4861 tgtggttgca tgagtgttct taacatactt cctgatgatc tcggcttctt cagaatctct
     4921 gtcaaccacc ttaatgtcag ttttgagctt ctcatagttg acatcgatgg gatccttgct
     4981 gctatcatca gaccctcccc tgagcagact gtaggccacc tcgatgtcca gcaggttgtc
     5041 aagcatttcc gccttggcct gcacactgtc tgcattgttc aggagcggag gcttcttcat
     5101 cccaaagtcg tgggggatca gggtgtaaaa gcgatttgag agatccagga tctgagagtc
     5161 gctgctgccc tgagacaccg cctgctggac ctcactgaag atggagtatg cggcctggat
     5221 ctgccttttg ctcagcttcc ccaagggcat cttctgaagg tcgatctcat actccaccat
     5281 ggctttcttc atactttcca catcaaagat catcttgatg aggtcctgaa ctggcttggg
     5341 gagcttggac ttggtgccag gatttactgt cagcttcttc actgcctctt catcctggcc
     5401 atagtcaatc tccagggggt agaacttttt gggatacttc gtgaaatttt tggagtgcca
     5461 agcgttcccg gttttttctt catataattt catgaagtgc tcaatggcat cctccttgga
     5521 cggcatctgt tccagtttgt tgctaccgat caccgtaccc acacggcccc aggacctgaa
     5581 tatccaatac ctgttttcct tgtcgtcctc cagaagctgg agcttgtagt aggagttggt
     5641 tcctttaacg atgtccacca ggccaagggt ggcactgaag accttcccac ctttctccag
     5701 gacatgcgca gagtgttcca gtccagaatc aggatccaca gctgctcctc ctttaagagt
     5761 taatttcatt ctcttttcag atttgttgat accttcctcc ttgacctggc ccttgctttt
     5821 tttggagagc gcagcccctg acttccctct tggggccaca acttcaacag gctctgcctt
     5881 cacctctgcc ccccaagggg acaagatgtg cgctaagaac aactcctgaa ggctcttggt
     5941 ggaggcggag acgtcctgga ggaagtcctc agacacaact cggatgttgg cttcctttac
     6001 ttcctccatc ttcttattca tcttttccac ctcctttttg gtgctgatgc acagggaagc
     6061 cttgttggcc gtccccgtca acttcccccc gagtttctca atcatggcct tcacttcatc
     6121 cttgttccgg gacagcttcc cgagagtcag gatcttcatg ttggataatg gcttatctgc
     6181 tgaagcagag gagttcacag cagcaggagc cgaggccacc tcatcgcctt ttctctttcc
     6241 ttcactcttg actcctggga gctgcttctt cagggcttcc ttatcctctg tagcaaggag
     6301 gctgaagccc ttgagctgac tcgcactgta ctcgggccgg aaacccagct cctccctgtt
     6361 cttgacaaag cagcctggat ggtaccagcg gtcaatcatg cctagctgtg gcttctccgg
     6421 gtccaccatc ttcttggaca ggcgcacctg gcccttttct atcttctcca tacacccctt
     6481 gcacgtactt ctgttggact tggcatactc tgctgcaaag tcacccagag tcttctctgc
     6541 cttgctacca attccatcct ggcctttgcc tgtcactcct ccagcttccg ctgtcttctt
     6601 gactttctgc tggtcatccc accgaagctc agagaaccca tccacctcaa cgtcagggtg
     6661 ccggatggag tggcccacct tccagaagca ggagaagtgg taccagtgtg ggacttttcc
     6721 atcaaacatg ggcgactgca ccatgatggc catccggagc gagtccttgg ggatgctctc
     6781 gctgcatttc ttgcaagagg cgcgcccgct cttggcgtac tcgactcgat acagcttatc
     6841 agaagactcc gccatcatat ggctgccgcg cggcaccagg ccgctgctgt gatgatgatg
     6901 atgatggctg ctgcccatgg tatatctcct tcttaaagtt aaacaaaatt atttctagag
     6961 gggaattgtt atccgctcac aattccccta tagtgagtcg tattaatttc gcgggatcga
     7021 gatctcgatc ctctacgccg gacgcatcgt ggccggcatc accggcgcca caggtgcggt
     7081 tgctggcgcc tatatcgccg acatcaccga tggggaagat cgggctcgcc acttcgggct
     7141 catgagcgct tgtttcggcg tgggtatggt ggcaggcccc gtggccgggg gactgttggg
     7201 cgccatctcc ttgcatgcac cattccttgc ggcggcggtg ctcaacggcc tcaacctact
     7261 actgggctgc ttcctaatgc aggagtcgca taagggagag cgtcgagatc ccggacacca
     7321 tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca
     7381 gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt
     7441 atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa
     7501 aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg
     7561 cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt
     7621 cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt
     7681 cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc
     7741 aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg
     7801 aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acaccca
//
