LOCUS       sequence-349905-        6399 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of recombinant fusion protein in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6399)
  AUTHORS   Sowa ST, Galera-Prat A, Wazir S, Alanen HI, Maksimainen MM, Lehtio L
  TITLE     A molecular toolbox for ADP-ribosyl binding proteins.
  JOURNAL   Cell Rep Methods. 2021 Dec 20;1(8):100121. doi: 
            10.1016/j.crmeth.2021.100121. Epub 2021 Nov 11.
  PUBMED    34786571
REFERENCE   2  (bases 1 to 6399)
  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: "Cell Rep 
            Methods."; date: "2021-12-20"; volume: "1(8):100121. doi"; pages: " 
            10.1016/j.crmeth.2021.100121. Epub 2021 Nov 11"
COMMENT     Sequence Label: XRCC1-c003
FEATURES             Location/Qualifiers
     source          1..6399
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             4..21
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             join(22..24,25..735)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIKA
                     HFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(64..85)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(325..344)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     672..693
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             760..780
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1176..1193
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1246..1264)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1260..1307
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1344..1799
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1431..1450)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1641..1662
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1891..2706)
                     /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(1891..2706)
                     /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     2614..2633
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2698..2717
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2828..3416
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3317..3336
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3570..3587
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3602..3741
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3667..3686)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3827..3849
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3843..4034)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4843..5925)
                     /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     5887..5906
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5926..6003)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6209..6228
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6312..6331
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6312..6330
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6331..6355
                     /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             6370..6392
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 atgcaccatc atcatcatca tgtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
       61 atcctggtcg agctggacgg cgacgtaaac ggccacaggt tcagcgtgtc cggcgagggc
      121 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg
      181 cccgtgccct ggcccaccct cgtgaccacc ctgacctggg gcgtgcagtg cttcgcccgc
      241 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
      301 caggagcgta ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag
      361 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac
      421 ggcaacatcc tggggcacaa gctggagtac aacgccatca gcgacaacgt ctatatcacc
      481 gccgacaagc agaagaacgg catcaaggcc cacttcaaga tccgccacaa catcgaggac
      541 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
      601 ctgctgcccg acaaccacta cctgagcacc cagtccaagc tgagcaaaga ccccaacgag
      661 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg
      721 gacgagctgt acaagtcttc tggtgtagat ctgggtaccg agaacctgta cttccaatcc
      781 atgggagaag gcaccgagcc cagacgaccc cgagctggcc cagaggagct ggggaagatc
      841 cttcagggtg tggtagtggt gctgagtggc ttccagaacc ccttccgctc cgagctgcga
      901 gataaggccc tagagcttgg ggccaagtat cggccagact ggacccggga cagcacgcac
      961 ctcatctgtg cctttgccaa cacccccaag tacagccagg tcctaggcct gggaggccgc
     1021 atcgtgcgta aggagtgggt gctggactgt caccgcatgc gtcggcggct gccctcccgg
     1081 aggtacctca tggcagggcc ataggacggt ctccagtaaa ggtggatacg gatccgaatt
     1141 cgagctccgt cgacaagctt gcggccgcac tcgagcacca ccaccaccac cactgagatc
     1201 cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac
     1261 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
     1321 ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     1381 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     1441 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     1501 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     1561 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     1621 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     1681 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     1741 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     1801 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     1861 tcaaatatgt atccgctcat gaattaattc ttagaaaaac tcatcgagca tcaaatggaa
     1921 ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa
     1981 tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc
     2041 gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt
     2101 atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagtttatg
     2161 catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc
     2221 atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata cgcgatcgct
     2281 gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc
     2341 atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttccc
     2401 ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt
     2461 cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt
     2521 ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa
     2581 tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa
     2641 atcagcatcc atgttggaat ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg
     2701 gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgacca
     2761 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     2821 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     2881 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     2941 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     3001 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     3061 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     3121 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     3181 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     3241 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     3301 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     3361 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     3421 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     3481 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     3541 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     3601 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac
     3661 tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta
     3721 cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     3781 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     3841 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca
     3901 gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt
     3961 ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt
     4021 tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata
     4081 ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta
     4141 ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc
     4201 actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag
     4261 cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc
     4321 agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt
     4381 ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta
     4441 aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat gcgcacccgt
     4501 ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt ggcgggacca
     4561 gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga caggccgatc
     4621 atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc tgccggcacc
     4681 tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat agtcatgccc
     4741 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     4801 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     4861 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     4921 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     4981 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     5041 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     5101 atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact cggtaatggc
     5161 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     5221 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     5281 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     5341 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
     5401 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
     5461 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
     5521 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
     5581 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
     5641 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
     5701 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
     5761 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
     5821 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
     5881 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
     5941 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     6001 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag cccagtagta
     6061 ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag atggcgccca
     6121 acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg ctcatgagcc
     6181 cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg ccagcaaccg
     6241 cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc gagatctcga
     6301 tcccgcgaaa ttaatacgac tcactatagg ggaattgtga gcggataaca attcccctct
     6361 agaaataatt ttgtttaact ttaagaagga gatatacat
//
