LOCUS       sequence-208251-        6138 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 6138)
  AUTHORS   Brzovic PS, Lissounov A, Christensen DE, Hoyt DW, Klevit RE
  TITLE     A UbcH5/ubiquitin noncovalent complex is required for processive 
            BRCA1-directed ubiquitination.
  JOURNAL   Mol Cell. 2006 Mar 17. 21(6):873-80.
  PUBMED    16543155
REFERENCE   2  (bases 1 to 6138)
  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: "Mol Cell. 
            2006 Mar 17. 21(6):873-80."
COMMENT     Sequence Label: BARD1 26-327 pET28a
FEATURES             Location/Qualifiers
     source          1..6138
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1079..1101
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1116..1140
                     /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(1140..1159)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1141..1159)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1243..1262)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1468..1545
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1546..2628
                     /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(1565..1584)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3437..3628
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3622..3644)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3730..3869
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3785..3804
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3884..3901)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4055..4643)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4135..4154)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4754..4773)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4765..5580
                     /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             4765..5580
                     /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(4838..4857)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5672..6127)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5809..5830)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6021..6040
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagttatcta ttgtgatggc
      181 cacgttttcc attattttct aatggcaaag atttcttaga tgtaagataa tttttgcaga
      241 ccttctcagg agtcactact tcattcctgc tcttagtgtc tggagactct atttgctcag
      301 ccaatggtaa agagacttca gttaaacttc caaaacattc agattctgtc aaggagccac
      361 ttgctagtaa gtctatttca ccatttatct gaggactgga gataacagat ggttggctac
      421 agaaggatac cagcttttgc ttagattcct ctttggagtc aaattcacca tcttcttttt
      481 ctgcctctaa attccatttt tggttgattt cagctaaagt tttctttttt tgcttttttc
      541 cagatcttgc agaagccttt ttagccctct cagaaacatc tgcaggagga cttggggaaa
      601 caaattcata tgagtcttgc tgagcacttg catctttttt tattgcaggc tgggtttgca
      661 ctgaagcttt actcacaaca tatctgactt ccttacttcg agggctaaac cacattttaa
      721 ttgaattctt cttgtttcct gcatcattaa acaaactttt cctaggttta tcttctttca
      781 aatctgacag ctcattgtca tgtagcaaat ttcgaagctt actacaaagt tgaatcatgc
      841 tgtccagttg tctatttatc ttcaagtctt gtatccaggc cggggtgtaa cacactggac
      901 atccagttcc aatgcagtca cttacacaat tactacagaa gatgtgctca catcctccta
      961 aacacacagg ctctctcaga atgttagtac aacgcgagca gcgcagcagc ttctccaggc
     1021 ggtcgagcgc ggcgcgactg tgggcccagg caccgcgacc atccggttcc atggtatatc
     1081 tccttcttaa agttaaacaa aattatttct agaggggaat tgttatccgc tcacaattcc
     1141 cctatagtga gtcgtattaa tttcgcggga tcgagatctc gatcctctac gccggacgca
     1201 tcgtggccgg catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca
     1261 ccgatgggga agatcgggct cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta
     1321 tggtggcagg ccccgtggcc gggggactgt tgggcgccat ctccttgcat gcaccattcc
     1381 ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt
     1441 cgcataaggg agagcgtcga gatcccggac accatcgaat ggcgcaaaac ctttcgcggt
     1501 atggcatgat agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg
     1561 ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac
     1621 caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg
     1681 aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc
     1741 gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct
     1801 cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa
     1861 gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac
     1921 tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg
     1981 ttatttcttg atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac
     2041 ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta
     2101 gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc
     2161 actcgcaatc aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt
     2221 tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc
     2281 aacgatcaga tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt
     2341 gcggatatct cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg
     2401 ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg
     2461 caactctctc agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa
     2521 agaaaaacca ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     2581 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     2641 taatgtaagt tagctcactc attaggcacc gggatctcga ccgatgccct tgagagcctt
     2701 caacccagtc agctccttcc ggtgggcgcg gggcatgact atcgtcgccg cacttatgac
     2761 tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca gcgctctggg tcattttcgg
     2821 cgaggaccgc tttcgctgga gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat
     2881 cttgcacgcc ctcgctcaag ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa
     2941 gcaggccatt atcgccggca tggcggcccc acgggtgcgc atgatcgtgc tcctgtcgtt
     3001 gaggacccgg ctaggctggc ggggttgcct tactggttag cagaatgaat caccgatacg
     3061 cgagcgaacg tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa caacatgaat
     3121 ggtcttcggt ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat
     3181 tatgttccgg atctgcatcg caggatgctg ctggctaccc tgtggaacac ctacatctgt
     3241 attaacgaag cgctggcatt gaccctgagt gatttttctc tggtcccgcc gcatccatac
     3301 cgccagttgt ttaccctcac aacgttccag taaccgggca tgttcatcat cagtaacccg
     3361 tatcgtgagc atcctctctc gtttcatcgg tatcattacc cccatgaaca gaaatccccc
     3421 ttacacggag gcatcagtga ccaaacagga aaaaaccgcc cttaacatgg cccgctttat
     3481 cagaagccag acattaacgc ttctggagaa actcaacgag ctggacgcgg atgaacaggc
     3541 agacatctgt gaatcgcttc acgaccacgc tgatgagctt taccgcagct gcctcgcgcg
     3601 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     3661 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     3721 gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac
     3781 tatgcggcat cagagcagat tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca
     3841 gatgcgtaag gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc
     3901 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     3961 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     4021 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     4081 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     4141 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4201 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     4261 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4321 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4381 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     4441 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     4501 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     4561 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     4621 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     4681 agtggaacga aaactcacgt taagggattt tggtcatgaa caataaaact gtctgcttac
     4741 ataaacagta atacaagggg tgttatgagc catattcaac gggaaacgtc ttgctctagg
     4801 ccgcgattaa attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat
     4861 gtcgggcaat caggtgcgac aatctatcga ttgtatggga agcccgatgc gccagagttg
     4921 tttctgaaac atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta
     4981 aactggctga cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat
     5041 gatgcatggt tactcaccac tgcgatcccc gggaaaacag cattccaggt attagaagaa
     5101 tatcctgatt caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat
     5161 tcgattcctg tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg
     5221 caatcacgaa tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc
     5281 tggcctgttg aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca
     5341 gtcgtcactc atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata
     5401 ggttgtattg atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta
     5461 tggaactgcc tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatggt
     5521 attgataatc ctgatatgaa taaattgcag tttcatttga tgctcgatga gtttttctaa
     5581 gaattaattc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     5641 tccgcgcaca tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa
     5701 ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa
     5761 atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac
     5821 aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag
     5881 ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt
     5941 aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg
     6001 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     6061 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     6121 ggcgcgtccc attcgcca
//
