LOCUS       sequence-9587-ex        6599 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6599)
  AUTHORS   Choi JH, Lindsey-Boltz LA, Sancar A
  TITLE     Cooperative activation of the ATR checkpoint kinase by TopBP1 and 
            damaged DNA.
  JOURNAL   Nucleic Acids Res. 2009 Apr;37(5):1501-9. doi: 10.1093/nar/gkn1075. 
            Epub 2009 Jan 12.
  PUBMED    19139065
REFERENCE   2  (bases 1 to 6599)
  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: 
            "10.1093/nar/gkn1075"; journalName: "Nucleic Acids Res"; date: 
            "2009-04"; volume: "37"; issue: "5"; pages: "1501-9"
COMMENT     Sequence Label: GST TopBP1 C (aa 978-1522)
FEATURES             Location/Qualifiers
     source          1..6599
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..941
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(1642..1653)
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     primer_bind     complement(2649..2671)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2816..2834)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2902..3006
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3007..3075,3076..3867)
                     /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(3225..3244)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4038..4626
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4527..4546
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4780..4797
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4870..4947
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             4948..6030
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4967..4986)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    6043..6064
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6079..6084,6085..6102,6103..6109)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6117..6133
                     /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     6141..6157
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6141..6157
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(6153..6599,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6173..6190)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6173..6189)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6182..6204)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gaagttctgt tccaggggcc cctgggatcc gcccaagagt
      961 gtaaacatct tcctgaatct ctttatccac atacttataa tcccaaaatg agcttggata
     1021 tcagcgcagt gcaagatggc cggctctgta atagtcgact actctcagct gtgtcttcaa
     1081 caaaggatga tgagccagat cctttgattt tagaagaaaa tgatgtagac aatatggcca
     1141 ccaataataa agagtcagca ccatcaaatg gaagtggaaa gaatgactct aaaggagttc
     1201 tgacacagac cttagagatg agagagaact ttcagaagca gttacaggag ataatgtctg
     1261 caacatcaat agtgaaaccc caagggcaga ggacttccct ttcaagaagt ggttgtaaca
     1321 gcgcatcttc aacccctgac agcactcgct ctgctcgcag tggacgaagt agagtcctag
     1381 aggcactgag gcagtctcgt cagacagtac ctgatgtcaa cacagagcct tcccaaaatg
     1441 aacagatcat ttgggatgac cctacagcaa gggaggagag agcaaggctt gccagcaatt
     1501 tgcagtggcc tagttgtccc acacaatact ctgagcttca ggttgacatt caaaacttgg
     1561 aggattctcc ttttcaaaag cctttacatg attcagaaat tgctaaacag gctgtctgtg
     1621 atcctggaaa catacgtgtg actgaagctc ccaaacaccc aatctctgaa gaactggaaa
     1681 ctcccataaa agacagccac ctgatcccta cgcctcaagc ccccagtatt gcctttccac
     1741 tcgccaaccc ccctgtggct ccgcacccta gagaaaagat tataacgata gaggagactc
     1801 atgaagaatt aaaaaaacag tacatatttc agttatcatc tctgaatcct caagaacgta
     1861 ttgactattg tcatctgatt gagaaactag gtggattggt gatagaaaag cagtgctttg
     1921 atcccacctg tacacacatt gttgtgggac atccacttcg aaacgagaag tatttagcct
     1981 cagtggcagc tgggaagtgg gtgcttcatc gctcctacct tgaagcctgc aggactgctg
     2041 gacacttcgt gcaggaagaa gactatgaat ggggaagtag ttccatactt gatgttctga
     2101 ctggaatcaa tgtacagcaa cgaagactag cacttgcagc aatgagatgg agaaaaaaaa
     2161 tccagcaaag acaagaatct ggcattgttg agggagcatt tagtgggtgg aaggttattt
     2221 tacatgtgga tcagtctcga gaagcaggct tcaaacgcct tcttcagtca ggaggagcaa
     2281 aggtgctacc tggtcattct gtacctttat ttaaagaggc cacacatctt ttttctgact
     2341 tgaataaact gaaaccagat gactcaggag ttaatatagc agaagctgct gcccagaacg
     2401 tgtactgctt gagaacagaa tacattgctg attatctcat gcaggaatca cctcctcatg
     2461 tagaaaatta ctgtctacca gaagctattt catttattca gaataataag gaacttggga
     2521 ctggattatc acaaaagagg aaagctccta cagaaaaaaa taaaatcaaa cgacctagag
     2581 tacactaagc ggccgcatcg tgactgactg acgatctgcc tcgcgcgttt cggtgatgac
     2641 ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat
     2701 gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca
     2761 gccatgaccc agtcacgtag cgatagcgga gtgtataatt cttgaagacg aaagggcctc
     2821 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     2881 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     2941 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     3001 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     3061 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     3121 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     3181 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     3241 ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     3301 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     3361 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     3421 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     3481 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     3541 acgatgcctg cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     3601 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     3661 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     3721 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     3781 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     3841 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     3901 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     3961 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     4021 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     4081 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     4141 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
     4201 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     4261 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     4321 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     4381 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     4441 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     4501 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     4561 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     4621 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     4681 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     4741 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     4801 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     4861 ataaattccg acaccatcga atggtgcaaa acctttcgcg gtatggcatg atagcgcccg
     4921 gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa cgttatacga tgtcgcagag
     4981 tatgccggtg tctcttatca gaccgtttcc cgcgtggtga accaggccag ccacgtttct
     5041 gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc tgaattacat tcccaaccgc
     5101 gtggcacaac aactggcggg caaacagtcg ttgctgattg gcgttgccac ctccagtctg
     5161 gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga tcaactgggt
     5221 gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa agcggcggtg
     5281 cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta actatccgct ggatgaccag
     5341 gatgccattg ctgtggaagc tgcctgcact aatgttccgg cgttatttct tgatgtctct
     5401 gaccagacac ccatcaacag tattattttc tcccatgaag acggtacgcg actgggcgtg
     5461 gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc attaagttct
     5521 gtctcggcgc gtctgcgtct ggctggctgg cataaatatc tcactcgcaa tcaaattcag
     5581 ccgatagcgg aacgggaagg cgactggagt gccatgtccg gttttcaaca aaccatgcaa
     5641 atgctgaatg agggcatcgt tcccactgcg atgctggttg ccaacgatca gatggcgctg
     5701 ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg
     5761 ggatacgacg ataccgaaga cagctcatgt tatatcccgc cgtcaaccac catcaaacag
     5821 gattttcgcc tgctggggca aaccagcgtg gaccgcttgc tgcaactctc tcagggccag
     5881 gcggtgaagg gcaatcagct gttgcccgtc tcactggtga aaagaaaaac caccctggcg
     5941 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     6001 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac
     6061 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
     6121 gagcggataa caatttcaca caggaaacag ctatgaccat gattacggat tcactggccg
     6181 tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag
     6241 cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
     6301 aacagttgcg cagcctgaat ggcgaatggc gctttgcctg gtttccggca ccagaagcgg
     6361 tgccggaaag ctggctggag tgcgatcttc ctgaggccga tactgtcgtc gtcccctcaa
     6421 actggcagat gcacggttac gatgcgccca tctacaccaa cgtaacctat cccattacgg
     6481 tcaatccgcc gtttgttccc acggagaatc cgacgggttg ttactcgctc acatttaatg
     6541 ttgatgaaag ctggctacag gaaggccaga cgcgaattat ttttgatggc gttggaatt
//
