LOCUS       sequence-342171-        6357 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of isolated PARP1 CAT domain containing L713F 
            gain-of-function mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6357)
  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 6357)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Proc Natl 
            Acad Sci U S A."; date: "2021-09-7"; volume: "118(36). pii: 
            2107277118. doi"; pages: " 10.1073/pnas.2107277118"
COMMENT     Sequence Label: pET28_6xHis-PARP1-CAT-L713F
FEATURES             Location/Qualifiers
     source          1..6357
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     544..563
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(583..660)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     866..885
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     969..988
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        969..987
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    988..1012
                     /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             1027..1049
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1068..1085
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1095..1112
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2190..2207
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2260..2278)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2274..2321
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2358..2813
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2445..2464)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2655..2676
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2905..3720)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2905..3720)
                     /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     3628..3647
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3712..3731
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3842..4430
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4331..4350
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4584..4601
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4616..4755
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4681..4700)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4841..4863
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4857..5048)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5857..6357,1..582))
                     /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"
ORIGIN
        1 cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac
       61 cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg
      121 tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc
      181 aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag
      241 aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac
      301 cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc
      361 gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg
      421 ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg
      481 cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac
      541 accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg
      601 actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc
      661 cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt
      721 tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca
      781 gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga
      841 agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac
      901 ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc
      961 cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt cccctctaga
     1021 aataattttg tttaacttta agaaggagat ataccatggg cagcagccat catcatcatc
     1081 atcacagcag cggcctggtg ccgcgcggca gccatatgac caagtccaag ctccccaagc
     1141 cagttcagga cctcatcaag atgatctttg atgtggaaag tatgaagaaa gccatggtgg
     1201 agtatgagat cgaccttcag aagatgccct tggggaagct gagcaaaagg cagatccagg
     1261 ccgcatactc catcttcagt gaggtccagc aggcggtgtc tcagggcagc agcgactctc
     1321 agatcctgga tctctcaaat cgcttttaca ccctgatccc ccacgacttt gggatgaaga
     1381 agcctccgct cctgaacaat gcagacagtg tgcaggccaa ggcggaaatg cttgacaacc
     1441 tgctggacat cgaggtggcc tacagtctgc tcaggggagg gtctgatgat agcagcaagg
     1501 atcccatcga tgtcaactat gagaagctca aaactgacat taaggtggtt gacagagatt
     1561 ctgaagaagc cgagatcatc aggaagtatg ttaagaacac tcatgcaacc acacacaatg
     1621 cgtatgactt ggaagtcatc gatatcttta agatagagcg tgaaggcgaa tgccagcgtt
     1681 acaagccctt taagcagctt cataaccgaa gattgctgtg gcacgggtcc aggaccacca
     1741 actttgctgg gatcctgtcc cagggtcttc ggatagcccc gcctgaagcg cccgtgacag
     1801 gctacatgtt tggtaaaggg atctatttcg ctgacatggt ctccaagagt gccaactact
     1861 gccatacgtc tcagggagac ccaataggct taatcctgtt gggagaagtt gcccttggaa
     1921 acatgtatga actgaagcac gcttcacata tcagcaagtt acccaagggc aagcacagtg
     1981 tcaaaggttt gggcaaaact acccctgatc cttcagctaa cattagtctg gatggtgtag
     2041 acgttcctct tgggaccggg atttcatctg gtgtgaatga cacctctcta ctatataacg
     2101 agtacattgt ctatgatatt gctcaggtaa atctgaagta tctgctgaaa ctgaaattca
     2161 attttaagac ctccctgtgg taactcgagc accaccacca ccaccactga gatccggctg
     2221 ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat
     2281 aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat
     2341 ccggattggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     2401 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     2461 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     2521 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     2581 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     2641 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     2701 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     2761 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc
     2821 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     2881 atgtatccgc tcatgaatta attcttagaa aaactcatcg agcatcaaat gaaactgcaa
     2941 tttattcata tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg
     3001 agaaaactca ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc
     3061 gactcgtcca acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag
     3121 tgagaaatca ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc
     3181 tttccagact tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac
     3241 caaaccgtta ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa
     3301 aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac
     3361 aatattttca cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat
     3421 cgcagtggtg agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag
     3481 aggcataaat tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac
     3541 gctacctttg ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata
     3601 gattgtcgca cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc
     3661 atccatgttg gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa tatggctcat
     3721 aacacccctt gtattactgt ttatgtaagc agacagtttt attgttcatg accaaaatcc
     3781 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     3841 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     3901 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     3961 tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact
     4021 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     4081 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     4141 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     4201 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     4261 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     4321 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     4381 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     4441 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     4501 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     4561 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga
     4621 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag
     4681 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
     4741 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     4801 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     4861 aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg
     4921 tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc
     4981 agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt
     5041 ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg
     5101 aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa
     5161 cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag
     5221 ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat
     5281 cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt
     5341 tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag
     5401 cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa
     5461 ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggggcc
     5521 gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg
     5581 aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc
     5641 gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct
     5701 acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc
     5761 caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc
     5821 taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     5881 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     5941 attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt
     6001 caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg
     6061 aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc
     6121 gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat
     6181 tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt
     6241 cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc
     6301 tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac gcagacg
//
