LOCUS       sequence-151486-        7868 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This plasmid encodes the Argonaute protein from Marinitoga 
            Piezophila and is codon optimized for E. coli expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7868)
  AUTHORS   Kaya E, Doxzen KW, Knoll KR, Wilson RC, Strutt SC, Kranzusch PJ, 
            Doudna JA
  TITLE     A bacterial Argonaute with noncanonical guide RNA specificity.
  JOURNAL   Proc Natl Acad Sci U S A. 2016 Mar 30. pii: 201524385.
  PUBMED    27035975
REFERENCE   2  (bases 1 to 7868)
  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: "Proc Natl 
            Acad Sci U S A. 2016 Mar 30. pii: 201524385."
COMMENT     Sequence Label: pET-6His-MBP-MpAgo
FEATURES             Location/Qualifiers
     source          1..7868
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(18..36)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        104..208
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(209..277,278..1069)
                     /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(427..446)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1240..1828
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1729..1748
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1982..1999
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2014..2156
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2082..2101)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2242..2264
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2258..2449)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     3907..3926
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     4010..4029
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4010..4028
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             4117..4139
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4159..4188
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             4198..5298
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     5272..5295
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     primer_bind     complement(7404..7422)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      7418..7465
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(7831..7836,7837..7853,7854..7859))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
ORIGIN
        1 ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
       61 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
      121 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
      181 gcttcaataa cattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
      241 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
      301 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
      361 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
      421 agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg
      481 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
      541 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
      601 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
      661 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
      721 accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact
      781 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
      841 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
      901 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
      961 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     1021 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     1081 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     1141 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     1201 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     1261 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     1321 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     1381 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     1441 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     1501 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     1561 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     1621 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     1681 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     1741 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     1801 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     1861 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     1921 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     1981 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     2041 tctgtgcggt atttcacacc gcatatatgg tgcactctca gtacaatctg ctctgatgcc
     2101 gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc
     2161 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     2221 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     2281 cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga
     2341 tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc
     2401 ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg
     2461 tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca
     2521 cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac
     2581 tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg
     2641 ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga
     2701 acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga
     2761 agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc
     2821 gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg
     2881 tcctcaacga caggagcacg atcatgcgca cccgtggcca ggacccaacg ctgcccgaga
     2941 tgcgccgcgt gcggctgctg gagatggcgg acgcgatgga tatgttctgc caagggttgg
     3001 tttgcgcatt cacagttctc cgcaagaatt gattggctcc aattcttgga gtggtgaatc
     3061 cgttagcgag gtgccgccgg cttccattca ggtcgaggtg gcccggctcc atgcaccgcg
     3121 acgcaacgcg gggaggcaga caaggtatag ggcggcgcct acaatccatg ccaacccgtt
     3181 ccatgtgctc gccgaggcgg cataaatcgc cgtgacgatc agcggtccag tgatcgaagt
     3241 taggctggta agagccgcga gcgatccttg aagctgtccc tgatggtcgt catctacctg
     3301 cctggacagc atggcctgca acgcgggcat cccgatgccg ccggaagcga gaagaatcat
     3361 aatggggaag gccatccagc ctcgcgtcgc gaacgccagc aagacgtagc ccagcgcgtc
     3421 ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt
     3481 gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat
     3541 cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg
     3601 tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg
     3661 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gacgctctcc
     3721 cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc
     3781 cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg ccacggggcc
     3841 tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc
     3901 cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc
     3961 ggccacgatg cgtccggcgt agaggatcga gatctcgatc ccgcgaaatt aatacgactc
     4021 actataggga gaccacaacg gtttccctct agtgccggct ccggagagct ctttaattaa
     4081 gcggccgccc tgcaggactc gagttctaga aatatttttg tttaacttta agaaggagat
     4141 atacatatga aatcttctca ccatcaccat caccatcacc atcaccatgg ttcttctatg
     4201 aaaatcgaag aaggtaaact ggtaatctgg attaacggcg ataaaggcta taacggtctc
     4261 gctgaagtcg gtaagaaatt cgagaaagat accggaatta aagtcaccgt tgagcatccg
     4321 gataaactgg aagagaaatt cccacaggtt gcggcaactg gcgatggccc tgacattatc
     4381 ttctgggcac acgaccgctt tggtggctac gctcaatctg gcctgttggc tgaaatcacc
     4441 ccggacaaag cgttccagga caagctgtat ccgtttacct gggatgccgt acgttacaac
     4501 ggcaagctga ttgcttaccc gatcgctgtt gaagcgttat cgctgattta taacaaagat
     4561 ctgctgccga acccgccaaa aacctgggaa gagatcccgg cgctggataa agaactgaaa
     4621 gcgaaaggta agagcgcgct gatgttcaac ctgcaagaac cgtacttcac ctggccgctg
     4681 attgctgctg acgggggtta tgcgttcaag tatgaaaacg gcaagtacga cattaaagac
     4741 gtgggcgtgg ataacgctgg cgcgaaagcg ggtctgacct tcctggttga cctgattaaa
     4801 aacaaacaca tgaatgcaga caccgattac tccatcgcag aagctgcctt taataaaggc
     4861 gaaacagcga tgaccatcaa cggcccgtgg gcatggtcca acatcgacac cagcaaagtg
     4921 aattatggtg taacggtact gccgaccttc aagggtcaac catccaaacc gttcgttggc
     4981 gtgctgagcg caggtattaa cgccgccagt ccgaacaaag agctggcaaa agagttcctc
     5041 gaaaactatc tgctgactga tgaaggtctg gaagcggtta ataaagacaa accgctgggt
     5101 gccgtagcgc tgaagtctta cgaggaagag ttggcgaaag atccacgtat tgccgccact
     5161 atggaaaacg cccagaaagg tgaaatcatg ccgaacatcc cgcagatgtc cgctttctgg
     5221 tatgccgtgc gtactgcggt gatcaacgcc gccagcggtc gtcagactgt cgatgaagcc
     5281 ctgaaagacg cgcagactaa ttcgagctcg gattacgata tcccaacgac cctagaagtg
     5341 ctttttcagg gcgccaatgc atacctgaat ctgtataaaa tcgacatccc gaagaaaatc
     5401 aaacgcttgt atttctataa tccggatatg gagccaaagc tcttcgcacg taatctgtcc
     5461 cgtgttaaca actttaagtt ccaggactcg aatgaccttg tgtggattga aatccctgac
     5521 atcgattttc aaatcacacc taaaaatgta tttcagtaca aagttgagaa agaagaaatt
     5581 atcaaagaag aagaagataa aaagctgttt gtcaagacac tgtataaata catcaagaag
     5641 ctgttcctcg ataacgattt ctattttaaa aaaggcaaca actttattag caactcggag
     5701 gtcttcagct tagatagcaa tgaaaatgta aacgcccatt taacttataa aatcaagatt
     5761 cataacattt cgaatgagta ctatctgtca attcttccga aatttacctt tctttcgaag
     5821 gaaccggcat tggaaagcgc aatcaaaagc ggttatttgt ataatatcaa gtcgggcaag
     5881 tctttcccat atattagcgg cctggacgga atcctgaaaa ttgacatcgg aaataatcag
     5941 atcgtggaag tggcgtatcc ggaaaattat ctgtttaact ttaccacgcg cgatgctgaa
     6001 aaatatggct ttagcaaaga agtacatgag atctataaaa acaaagtctt cgaaggcttt
     6061 aaaaaaatcc cgaagactct gggcttctta aataaaatta caaatctgaa cgagaattac
     6121 cagttgaaag acggctacaa aatcttcatt aacgtgatct ataaattcaa aaacggcgag
     6181 agccgttatg ccaaagatgt gtttaaatac tccttttata aaaatgaaca gccattaaaa
     6241 gccatcttct ttttctcttc gaaaaaacaa ttttttgaag tgcagaaatc gcttaaagaa
     6301 ctgtttcata acaaacactc ggtgttctat cgcgccgccg cggagctggg cttctctaaa
     6361 gtcgaatttt tacgtgatag caaaacgaaa tcctctgcct tcctgtacaa tccggaggag
     6421 ttcaccgtga aaaatacgga atttattaac caaatcgaag ataatgttat ggccatcgtc
     6481 ttactggaca aatacattgg taatatcgat ccgctcgtac gtaattttcc agataattta
     6541 atccttcaac cgatcctgaa agaaaaattg gaagatatta aaccttttat catcaaatca
     6601 tatgtgtata aaatgggaaa ctttatccca gaatgcaagc ctttcatcct gaaaaaaatg
     6661 gaagataaag agaaaaactt gtatattggg attgatcttt cccatgatac gtacgcacgt
     6721 aaaactaacc tgtgtattgc ggccgttgat aacaccggcg atatcttgta tattggcaaa
     6781 cataaaaacc tggaattgaa cgaaaagatg aatttggata ttctcgaaaa agaatatatt
     6841 aaggcttttg aaaagtatat tgagaaattc aacgtgtccc cggaaaatgt atttattttg
     6901 cgcgatggcc ggtttatcga agacattgag attattaaaa atttcatctc atataatgat
     6961 actaagtata cactggtgga agtaaataaa aatacgaaca ttaatagtta tgacgatctc
     7021 aaagaatgga ttattaaact ggacgaaaac acctatatct attacccgaa aacttttctg
     7081 aatcaaaaag gggttgaagt taaaatcctg gagaacaaca ccgattatac gattgaagaa
     7141 atcattgagc agatctatct gctgacccgt gtggcgcaca gcaccccgta cactaactat
     7201 aaattgccgt acccgctcca catcgcaaat aaggtggcgt tgacggatta cgagtggaag
     7261 ctgtatattc cgtactaata acattggaag tggataacgg atccgcgatc gcggcgcgcc
     7321 acctggtggc cggccggtac cacgcgtgcg cgctgatccg gctgctaaca aagcccgaaa
     7381 ggaagctgag ttggctgctg ccaccgctga gcaataacta gcataacccc ttggggcctc
     7441 taaacgggtc ttgaggggtt ttttgctgaa aggaggaact atatccggat atccacagga
     7501 cgggtgtggt cgccatgatc gcgtagtcga tagtggctcc aagtagcgaa gcgagcagga
     7561 ctgggcggcg gccaaagcgg tcggacagtg ctccgagaac gggtgcgcat agaaattgca
     7621 tcaacgcata tagcgctagc agcacgccat agtgactggc gatgctgtcg gaatggacga
     7681 tatcccgcaa gaggcccggc agtaccggca taaccaagcc tatgcctaca gcatccaggg
     7741 tgacggtgcc gaggatgacg atgagcgcat tgttagattt catacacggt gcctgactgc
     7801 gttagcaatt taactgtgat aaactaccgc attaaagctt atcgatgata agctgtcaaa
     7861 catgagaa
//
