LOCUS       sequence-464128-        6401 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of exodeoxyribonuclease III 
            (Exonuclease III) in Escherichia coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6401)
  TITLE     AI Proteins, Inc. plasmids
REFERENCE   2  (bases 1 to 6401)
  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"
COMMENT     Sequence Label: pAIP4_1AKO
FEATURES             Location/Qualifiers
     source          1..6401
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             547..738
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(732..754)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    840..979
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     895..914
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(994..1011)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1165..1753)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1245..1264)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(1864..1883)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1875..2690
                     /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             1875..2690
                     /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(1948..1967)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(2782..3237)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2919..2940)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3131..3150
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(3274..3321)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     3317..3335
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(3388..3405)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(4534..4563)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             4586..4608
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    4623..4647
                     /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(4647..4666)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4648..4666)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4754..4773)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        4979..5056
                     /gene="lacI"
                     /label=lacI promoter
     CDS             5057..6139
                     /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(5076..5095)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ctcgctcaag ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa gcaggccatt
       61 atcgccggca tggcggcccc acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg
      121 ctaggctggc ggggttgcct tactggttag cagaatgaat caccgatacg cgagcgaacg
      181 tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa caacatgaat ggtcttcggt
      241 ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg
      301 atctgcatcg caggatgctg ctggctaccc tgtggaacac ctacatctgt attaacgaag
      361 cgctggcatt gaccctgagt gatttttctc tggtcccgcc gcatccatac cgccagttgt
      421 ttaccctcac aacgttccag taaccgggca tgttcatcat cagtaacccg tatcgtgagc
      481 atcctctctc gtttcatcgg tatcattacc cccatgaaca gaaatccccc ttacacggag
      541 gcatcagtga ccaaacagga aaaaaccgcc cttaacatgg cccgctttat cagaagccag
      601 acattaacgc ttctggagaa actcaacgag ctggacgcgg atgaacaggc agacatctgt
      661 gaatcgcttc acgaccacgc tgatgagctt taccgcagct gcctcgcgcg tttcggtgat
      721 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
      781 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
      841 gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat
      901 cagagcagat tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca gatgcgtaag
      961 gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     1021 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     1081 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     1141 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     1201 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     1261 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     1321 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     1381 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     1441 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     1501 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     1561 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     1621 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     1681 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     1741 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     1801 aaactcacgt taagggattt tggtcatgaa caataaaact gtctgcttac ataaacagta
     1861 atacaagggg tgttatgagc catattcaac gggaaacgtc ttgctctagg ccgcgattaa
     1921 attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat
     1981 caggtgcgac aatctatcga ttgtatggga agcccgatgc gccagagttg tttctgaaac
     2041 atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga
     2101 cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt
     2161 tactcaccac tgcgatcccc gggaaaacag cattccaggt attagaagaa tatcctgatt
     2221 caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg
     2281 tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa
     2341 tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg
     2401 aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca gtcgtcactc
     2461 atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg
     2521 atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc
     2581 tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc
     2641 ctgatatgaa taaattgcag tttcatttga tgctcgatga gtttttctaa gaattaattc
     2701 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     2761 tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa
     2821 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
     2881 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
     2941 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
     3001 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
     3061 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
     3121 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
     3181 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc
     3241 attcgccaat ccggatatag ttcctccttt cagcaaaaaa cccctcaaga cccgtttaga
     3301 ggccccaagg ggttatgcta gttattgctc agcggtggca gcagccaact cagcttcctt
     3361 tcgggctttg ttagcagccg gatctcagtg gtggtggtgg tggtgctcga gtcattacga
     3421 accacgccta aaggtagccc aaaccggcgc gtgatccgac ggcttttcca tagaacggat
     3481 ctcatagtca ataccggttt cgacacagca ctctgcgagc ggctggcttg caagcagcag
     3541 gtcgatgcgc agaccacgat tgtcgtcgaa gcccttgcta cgataatcaa accagctaaa
     3601 acgatccgcg gtttgtggat tggcgtggcg gaacgtatca accagacccc agctcattaa
     3661 gcggtccatc cattcgcgct cttctggcag gaagctgcat ttgccggtgc gtagccagcg
     3721 cttgcgattt tcctcgccaa ttccgatatc caaatcggtc ggagaaatat tcatatcgcc
     3781 catgatcagc accgggttgt ctcttttcag ttccgtttcc aggtagtttt gcaagttctg
     3841 gtaaaattgc gccttcgccg ggaatttaat cgggtggtca cgggattcac cctgcgggaa
     3901 gtagccgtta atcacggtca cgttgcccag cagggacggg atctctgcca tgatgatacg
     3961 acgctgagcc tcttcgtcgt cgccagggaa accacgacgt acggcaatcg gagtctcttt
     4021 ggtcaacagc gccacaccgt agtgaccctt ctgaccatga taaaagacgt tgtaacccag
     4081 cttcgcaacc tcctccagcg ggaacatgtc atcgtgaact ttggtttctt gcaagccaat
     4141 aacatccggt tgatgttttt ccacgatcgc ttccagctga tgcggacgcg cacgtaatcc
     4201 gtttatatta aatgatacga atttcattcc agagccgcca atctgttcac ggtgggcttc
     4261 aatgatgtcg ttgtcttcca tgtccaggtc ttccggcgct tgatctgcct ggatgcgaat
     4321 accgtcatac aggaaacgca ggctatccat ttccttgccc tgacgtttgg caaacgcttc
     4381 catcaggcga cgcagcggcg tggtcttttt gatcttaaag aaaatttcag agctgccgtc
     4441 actaactttc aggttaatat gggtttccgg tttcacttcc ggcttgactt ccggctttgc
     4501 ttcttggttg acttccgaat cttgcaggct accgtggtga tggtgatggt gatgatggtg
     4561 atgaatcttc gccatcatat gtatatctcc ttcttaaagt taaacaaaat tatttctaga
     4621 ggggaattgt tatccgctca caattcccct atagtgagtc gtattaattt cgcgggatcg
     4681 agatcgatct cgatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt
     4741 gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc
     4801 gggctcatga gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg
     4861 ttgggcgcca tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac
     4921 ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgtcg agatcccgga
     4981 caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca
     5041 attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt
     5101 ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg
     5161 ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca
     5221 actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc
     5281 gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt
     5341 ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct
     5401 cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc
     5461 tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc
     5521 catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt
     5581 cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg
     5641 tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga
     5701 acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga
     5761 gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg
     5821 cgccattacc gagtccgggc tgcgcgttgg tgcggacatc tcggtagtgg gatacgacga
     5881 taccgaagac agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct
     5941 gctggggcaa accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg
     6001 caatcagctg ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca
     6061 aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg
     6121 actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac
     6181 cgggatctcg accgatgccc ttgagagcct tcaacccagt cagctccttc cggtgggcgc
     6241 ggggcatgac tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac
     6301 aggtgccggc agcgctctgg gtcattttcg gcgaggaccg ctttcgctgg agcgcgacga
     6361 tgatcggcct gtcgcttgcg gtattcggaa tcttgcacgc c
//
