LOCUS       sequence-350560-        5993 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial vector for expression of an N-terminal GST fusion of 
            PolB(K206A/K244A/T304I) with a TEV protease site located between the
            GST tag and PolB(K206A/K244A/T304I).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5993)
  AUTHORS   Fang Q, Andrews J, Sharma N, Wilk A, Clark J, Slyskova J, Koczor CA,
            Lans H, Prakash A, Sobol RW
  TITLE     Stability and sub-cellular localization of DNA polymerase beta is 
            regulated by interactions with NQO1 and XRCC1 in response to 
            oxidative stress.
  JOURNAL   Nucleic Acids Res. 2019 Jul 9;47(12):6269-6286. doi: 
            10.1093/nar/gkz293.
  PUBMED    31287140
REFERENCE   2  (bases 1 to 5993)
  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"; doi: "10"; journalName: 
            "Nucleic Acids Res"; date: "2019-07-9- 9"; volume: "47"; issue: 
            "12"; pages: "6269-6286"
COMMENT     Sequence Label: pGEX4T3-GST-PolB(K206A/K244A/T304I)
FEATURES             Location/Qualifiers
     source          1..5993
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(175..180,181..198,199..205)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    213..229
                     /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     237..253
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     237..253
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             249..776
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(269..286)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(269..285)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(278..300)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(878..883,884..899,900..906)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    914..930
                     /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)."
     CDS             953..1606
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1564..1586
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1613..1630
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1631..1651
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(2738..2760)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2905..2923)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2991..3095
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3096..3164,3165..3956)
                     /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(3314..3333)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4127..4715
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4616..4635
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4869..4886
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4959..5036
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(5037..5993,1..126)
                     /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(5056..5075)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac cgcctctccc
       61 cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg
      121 cagtgagcgc aacgcaatta atgtaagtta gctcactcat taggcacccc aggctttaca
      181 ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg
      241 aaacagctat gaccatgatt acggattcac tggccgtcgt tttacaacgt cgtgactggg
      301 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
      361 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
      421 aatggcgctt tgcctggttt ccggcaccag aagcggtgcc ggaaagctgg ctggagtgcg
      481 atcttcctga ggccgatact gtcgtcgtcc cctcaaactg gcagatgcac ggttacgatg
      541 cgcccatcta caccaacgtg acctatccca ttacggtcaa tccgccgttt gttcccacgg
      601 agaatccgac gggttgttac tcgctcacat ttaatgttga tgaaagctgg ctacaggaag
      661 gccagacgcg aattattttt gatggcgttg gaattagctt atcgactgca cggtgcacca
      721 atgcttctgg cgtcaggcag ccatcggaag ctgtggtatg gctgtgcagg tcgtaaatca
      781 ctgcataatt cgtgtcgctc aaggcgcact cccgttctgg ataatgtttt ttgcgccgac
      841 atcataacgg ttctggcaaa tattctgaaa tgagctgttg acaattaatc atcggctcgt
      901 ataatgtgtg gaattgtgag cggataacaa tttcacacag gaaacagtat tcatgtcccc
      961 tatactaggt tattggaaaa ttaagggcct tgtgcaaccc actcgacttc ttttggaata
     1021 tcttgaagaa aaatatgaag agcatttgta tgagcgcgat gaaggtgata aatggcgaaa
     1081 caaaaagttt gaattgggtt tggagtttcc caatcttcct tattatattg atggtgatgt
     1141 taaattaaca cagtctatgg ccatcatacg ttatatagct gacaagcaca acatgttggg
     1201 tggttgtcca aaagagcgtg cagagatttc aatgcttgaa ggagcggttt tggatattag
     1261 atacggtgtt tcgagaattg catatagtaa agactttgaa actctcaaag ttgattttct
     1321 tagcaagcta cctgaaatgc tgaaaatgtt cgaagatcgt ttatgtcata aaacatattt
     1381 aaatggtgat catgtaaccc atcctgactt catgttgtat gacgctcttg atgttgtttt
     1441 atacatggac ccaatgtgcc tggatgcgtt cccaaaatta gtttgtttta aaaaacgtat
     1501 tgaagctatc ccacaaattg ataagtactt gaaatccagc aagtatatag catggccttt
     1561 gcagggctgg caagccacgt ttggtggtgg cgaccatcct ccaaaatcgg atctggttcc
     1621 gcgtggatcc gaaaacctgt attttcaggg ctcgaattcc cgggtcgaca tgagcaaacg
     1681 gaaggcgccg caggagactc tcaacggggg aatcaccgac atgctcacag aactcgcaaa
     1741 ctttgagaag aacgtgagcc aagctatcca caagtacaat gcttacagaa aagcagcatc
     1801 tgttatagca aaatacccac acaaaataaa gagtggagct gaagctaaga aattgcctgg
     1861 agtaggaaca aaaattgctg aaaagattga tgagttttta gcaactggaa aattacgtaa
     1921 actggaaaag attcggcagg atgatacgag ttcatccatc aatttcctga ctcgagttag
     1981 tggcattggt ccatctgctg caaggaagtt tgtagatgaa ggaattaaaa cactagaaga
     2041 tctcagaaaa aatgaagata aattgaacca tcatcagcga attgggctga aatattttgg
     2101 ggactttgaa aaaagaattc ctcgtgaaga gatgttacaa atgcaagata ttgtactaaa
     2161 tgaagttaaa aaagtggatt ctgaatacat tgctacagtc tgtggcagtt tcagaagagg
     2221 tgcagagtcc agtggtgaca tggatgttct cctgacccat cccagcttca cttcagaatc
     2281 aaccgcacag ccaaaactgt tacatcaggt tgtggagcag ttacaaaagg ttcattttat
     2341 cacagatacc ctgtcaaagg gtgagacaaa gttcatgggt gtttgccagc ttcccagtgc
     2401 aaatgatgaa aaagaatatc cacacagaag aattgatatc aggttgatac ccaaagatca
     2461 gtattactgt ggtgttctct atttcactgg gagtgatatt ttcaataaga atatgagggc
     2521 tcatgcccta gaaaagggtt tcacaatcaa tgagtacacc atccgtccct tgggagtcat
     2581 tggagttgca ggagaacccc tgccagtgga tagtgaaaaa gacatctttg attacatcca
     2641 gtggaaatac cgggaaccca aggaccggag cgaatgagcg gccgcatcgt gactgactga
     2701 cgatctgcct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     2761 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     2821 cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag
     2881 tgtataattc ttgaagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     2941 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     3001 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     3061 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     3121 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     3181 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     3241 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     3301 cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg caagagcaac
     3361 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     3421 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     3481 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     3541 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     3601 aagccatacc aaacgacgag cgtgacacca cgatgcctgc agcaatggca acaacgttgc
     3661 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     3721 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     3781 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     3841 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     3901 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     3961 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     4021 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     4081 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     4141 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     4201 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     4261 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     4321 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     4381 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     4441 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     4501 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     4561 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     4621 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     4681 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     4741 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     4801 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     4861 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc
     4921 ttacgcatct gtgcggtatt tcacaccgca taaattccga caccatcgaa tggcgcaaaa
     4981 cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg gtgaatgtga
     5041 aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag accgtttccc
     5101 gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga
     5161 tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc aaacagtcgt
     5221 tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg
     5281 cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa
     5341 gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc
     5401 tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct gcctgcacta
     5461 atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt attattttct
     5521 cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt caccagcaaa
     5581 tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg gctggctggc
     5641 ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc gactggagtg
     5701 ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt cccactgcga
     5761 tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc gagtccgggc
     5821 tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac agctcatgtt
     5881 atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa accagcgtgg
     5941 accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg ttg
//
