LOCUS       sequence-200667-        6088 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  bacterial expression of Exonuclease I.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6088)
  AUTHORS   Korada SK, Johns TD, Smith CE, Jones ND, McCabe KA, Bell CE
  TITLE     Crystal structures of Escherichia coli exonuclease I in complex with
            single-stranded DNA provide insights into the mechanism of 
            processive digestion.
  JOURNAL   Nucleic Acids Res. 2013 Jun;41(11):5887-97. doi: 10.1093/nar/gkt278.
            Epub 2013 Apr 22.
  PUBMED    23609540
REFERENCE   2  (bases 1 to 6088)
  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/gkt278"; journalName: "Nucleic Acids Res"; date: 
            "2013-06"; volume: "41"; issue: "11"; pages: "5887-97"
COMMENT     Sequence Label: pET14b – E. coli Exonuclease I
FEATURES             Location/Qualifiers
     source          1..6088
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             51..73
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             92..109
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             119..136
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1618..1636)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1632..1679
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(2045..2050,2051..2067,2068..2073))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(2100..2118)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2186..2290
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2291..2359,2360..3151)
                     /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(2509..2528)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3322..3910
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3811..3830
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4064..4081
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4096..4235
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4161..4180)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4321..4343
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4337..4528)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     5986..6005
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg agaccacaac ggtttccctc tagaaataat tttgtttaac
       61 tttaagaagg agatatacca tgggcagcag ccatcatcat catcatcaca gcagcggcct
      121 ggtgccgcgc ggcagccata tgatgaatga cggtaagcaa caatctacct ttttgtttca
      181 cgattacgaa acctttggca cgcaccccgc gttagatcgc cctgcacagt tcgcagccat
      241 tcgcaccgat agcgaattca atgtcatcgg cgaacccgaa gtcttttact gcaagcccgc
      301 tgatgactat ttaccccagc caggagccgt attaattacc ggtattaccc cgcaggaagc
      361 acgggcgaaa ggagaaaacg aagccgcgtt tgccgcccgt attcactcgc tttttaccgt
      421 accgaagacc tgtattctgg gctacaacaa tgtgcgtttc gacgacgaag tcacacgcaa
      481 cattttttat cgtaatttct acgatcctta cgcctggagc tggcagcatg ataactcgcg
      541 ctgggattta ctggatgtta tgcgtgcctg ttatgccctg cgcccggaag gaataaactg
      601 gcctgaaaat gatgacggtc taccgagctt tcgccttgag catttaacca aagcgaatgg
      661 tattgaacat agcaacgccc acgatgcgat ggctgatgtg tacgccacta ttgcgatggc
      721 aaagctggta aaaacgcgtc agccacgcct gtttgattat ctctttaccc atcgtaataa
      781 acacaaactg atggcgttga ttgatgttcc gcagatgaaa cccctggtgc acgtttccgg
      841 aatgtttgga gcatggcgcg gcaataccag ctgggtggca ccgctggcgt ggcatcctga
      901 aaatcgcaat gccgtaatta tggtggattt ggcaggagac atttcgccat tactggaact
      961 ggatagcgac acattgcgcg agcgtttata taccgcaaaa accgatcttg gcgataacgc
     1021 cgccgttccg gttaagctgg tgcatatcaa taaatgtccg gtgctggccc aggcgaatac
     1081 gctacgcccg gaagatgccg accgactggg aattaatcgt cagcattgcc tcgataacct
     1141 gaaaattctg cgtgaaaatc cgcaagtgcg cgaaaaagtg gtggcgatat tcgcggaagc
     1201 cgaaccgttt acgccttcag ataacgtgga tgcacagctt tataacggct ttttcagtga
     1261 cgcagatcgt gcagcaatga aaattgtgct ggaaaccgag ccgcgtaatt taccggcact
     1321 ggatatcact tttgttgata aacggattga aaagctgttg ttcaattatc gggcacgcaa
     1381 cttcccgggg acgctggatt atgccgagca gcaacgctgg ctggagcacc gtcgccaggt
     1441 cttcacgcca gagtttttgc agggttatgc tgatgaattg cagatgctgg tacaacaata
     1501 tgccgatgac aaagagaaag tggcgctgtt aaaagcactt tggcagtacg cggaagagat
     1561 tgtctaagga tccggctgct aacaaagccc gaaaggaagc tgagttggct gctgccaccg
     1621 ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc
     1681 tgaaaggagg aactatatcc ggatatccac aggacgggtg tggtcgccat gatcgcgtag
     1741 tcgatagtgg ctccaagtag cgaagcgagc aggactgggc ggcggccaaa gcggtcggac
     1801 agtgctccga gaacgggtgc gcatagaaat tgcatcaacg catatagcgc tagcagcacg
     1861 ccatagtgac tggcgatgct gtcggaatgg acgatatccc gcaagaggcc cggcagtacc
     1921 ggcataacca agcctatgcc tacagcatcc agggtgacgg tgccgaggat gacgatgagc
     1981 gcattgttag atttcataca cggtgcctga ctgcgttagc aatttaactg tgataaacta
     2041 ccgcattaaa gcttatcgat gataagctgt caaacatgag aattcttgaa gacgaaaggg
     2101 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc
     2161 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     2221 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     2281 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     2341 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     2401 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     2461 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     2521 ggtattatcc cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     2581 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     2641 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     2701 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     2761 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     2821 caccacgatg cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     2881 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     2941 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     3001 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     3061 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     3121 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     3181 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     3241 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     3301 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     3361 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     3421 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta
     3481 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     3541 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     3601 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     3661 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     3721 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     3781 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     3841 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     3901 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     3961 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     4021 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     4081 ggaagcggaa gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
     4141 ccgcaatggt gcactctcag tacaatctgc tctgatgccg catagttaag ccagtataca
     4201 ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg acacccgcca acacccgctg
     4261 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     4321 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc
     4381 ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt
     4441 ccagctcgtt gagtttctcc agaagcgtta atgtctggct tctgataaag cgggccatgt
     4501 taagggcggt tttttcctgt ttggtcactg atgcctccgt gtaaggggga tttctgttca
     4561 tgggggtaat gataccgatg aaacgagaga ggatgctcac gatacgggtt actgatgatg
     4621 aacatgcccg gttactggaa cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg
     4681 accagagaaa aatcactcag ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc
     4741 cacagggtag ccagcagcat cctgcgatgc agatccggaa cataatggtg cagggcgctg
     4801 acttccgcgt ttccagactt tacgaaacac ggaaaccgaa gaccattcat gttgttgctc
     4861 aggtcgcaga cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat
     4921 tctgctaacc agtaaggcaa ccccgccagc ctagccgggt cctcaacgac aggagcacga
     4981 tcatgcgcac ccgtggccag gacccaacgc tgcccgagat gcgccgcgtg cggctgctgg
     5041 agatggcgga cgcgatggat atgttctgcc aagggttggt ttgcgcattc acagttctcc
     5101 gcaagaattg attggctcca attcttggag tggtgaatcc gttagcgagg tgccgccggc
     5161 ttccattcag gtcgaggtgg cccggctcca tgcaccgcga cgcaacgcgg ggaggcagac
     5221 aaggtatagg gcggcgccta caatccatgc caacccgttc catgtgctcg ccgaggcggc
     5281 ataaatcgcc gtgacgatca gcggtccaat gatcgaagtt aggctggtaa gagccgcgag
     5341 cgatccttga agctgtccct gatggtcgtc atctacctgc ctggacagca tggcctgcaa
     5401 cgcgggcatc ccgatgccgc cggaagcgag aagaatcata atggggaagg ccatccagcc
     5461 tcgcgtcgcg aacgccagca agacgtagcc cagcgcgtcg gccgccatgc cggcgataat
     5521 ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt gagcgagggc
     5581 gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc agcgaaagcg
     5641 gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt gcatgataaa
     5701 gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga aggagctgac
     5761 tgggttgaag gctctcaagg gcatcggtcg acgctctccc ttatgcgact cctgcattag
     5821 gaagcagccc agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg
     5881 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
     5941 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
     6001 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
     6061 gaggatcgag atctcgatcc cgcgaaat
//
