LOCUS       sequence-500739-        7091 bp    DNA     circular SYN 23-FEB-2026
DEFINITION  His-MBP-HamM from P. fluorescens for overexpression. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7091)
  AUTHORS   Doudna J et al.
  TITLE     Recurrent acquisition of nuclease-protease pairs in antiviral 
            immunity
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7091)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Feb 23, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: exp_PflHamM
FEATURES             Location/Qualifiers
     source          1..7091
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2..21
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2..20
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             109..131
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             151..180
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             190..1290
                     /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     1264..1287
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1345..1365
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     primer_bind     complement(2622..2640)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2636..2683
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(3049..3054,3055..3071,3072..3077))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(3104..3122)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3190..3294
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3295..3363,3364..4155)
                     /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(3513..3532)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4326..4914
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4815..4834
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5068..5085
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5100..5239
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5165..5184)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5325..5347
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5341..5532)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     6990..7009
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 ttaatacgac tcactatagg gagaccacaa cggtttccct ctagtgccgg ctccggagag
       61 ctctttaatt aagcggccgc cctgcaggac tcgagttcta gaaataattt tgtttaactt
      121 taagaaggag atatacatat gaaatcttct caccatcacc atcaccatca ccatcaccat
      181 ggttcttcta tgaaaatcga agaaggtaaa ctggtaatct ggattaacgg cgataaaggc
      241 tataacggtc tcgctgaagt cggtaagaaa ttcgagaaag ataccggaat taaagtcacc
      301 gttgagcatc cggataaact ggaagagaaa ttcccacagg ttgcggcaac tggcgatggc
      361 cctgacatta tcttctgggc acacgaccgc tttggtggct acgctcaatc tggcctgttg
      421 gctgaaatca ccccggacaa agcgttccag gacaagctgt atccgtttac ctgggatgcc
      481 gtacgttaca acggcaagct gattgcttac ccgatcgctg ttgaagcgtt atcgctgatt
      541 tataacaaag atctgctgcc gaacccgcca aaaacctggg aagagatccc ggcgctggat
      601 aaagaactga aagcgaaagg taagagcgcg ctgatgttca acctgcaaga accgtacttc
      661 acctggccgc tgattgctgc tgacgggggt tatgcgttca agtatgaaaa cggcaagtac
      721 gacattaaag acgtgggcgt ggataacgct ggcgcgaaag cgggtctgac cttcctggtt
      781 gacctgatta aaaacaaaca catgaatgca gacaccgatt actccatcgc agaagctgcc
      841 tttaataaag gcgaaacagc gatgaccatc aacggcccgt gggcatggtc caacatcgac
      901 accagcaaag tgaattatgg tgtaacggta ctgccgacct tcaagggtca accatccaaa
      961 ccgttcgttg gcgtgctgag cgcaggtatt aacgccgcca gtccgaacaa agagctggca
     1021 aaagagttcc tcgaaaacta tctgctgact gatgaaggtc tggaagcggt taataaagac
     1081 aaaccgctgg gtgccgtagc gctgaagtct tacgaggaag agttggcgaa agatccacgt
     1141 attgccgcca ctatggaaaa cgcccagaaa ggtgaaatca tgccgaacat cccgcagatg
     1201 tccgctttct ggtatgccgt gcgtactgcg gtgatcaacg ccgccagcgg tcgtcagact
     1261 gtcgatgaag ccctgaaaga cgcgcagact aattcgagct cgaacaacaa caacaataac
     1321 aataacaaca acctcgggat cgaggaaaac ctgtacttcc aatccaatgc aatgaaaatt
     1381 tccatgcacg atgccgcata cggtgattca attctcatca gcggcgcctc gaccaaaatt
     1441 ttagtggatg gtggaactgc gaagtccttc gctgactggt cgccttcgtt agagggtgtc
     1501 gatacgctag atgccctgat agttactcat attgataacg accatattgg cggagcaatc
     1561 aagctgctca catcaaagca cgcaactaaa gtaaaagaag tttatttcaa tggcattaac
     1621 caaatcctag atgtggagga ttgtgatatt ggtgatcgta gtgacgatct agaattatcc
     1681 gcaatcatag caaacaccac gtctagagac gaagaccagt tgatagggta ttctgaaggc
     1741 acatcgttat ccttaatatt acagcagcaa aatatcccct taaataaagc aaccgaaggc
     1801 aggcaaatcc acagaacgag ccttaataaa aacataataa taggagaatt tgaaattgaa
     1861 ctgataggcc caagcagaaa gagcttgtta aaaatccaac aaaaatggct tgaactatta
     1921 gaaagtaaag aaatcaaaag acgccttatc agcccaagat atagcgttgc attcgaaagc
     1981 tttatcagct cgattgatac ctcagtaaag catgatgagg aaatttcaag tctaagtgaa
     2041 aacatagagc atttggcaaa tcaaccattt actaatgaca catcaccccc aaacaaaagc
     2101 agcatctctt tccttctaaa aactgaaaac aaaagcattt taatgctcgg agattccgac
     2161 gcagagacta tagaggcatg gttgaactcg aagtctgcag atagcatcac agttgatgcg
     2221 gtaaagctac cccatcacgg aagccgaaat aactttagtg ctaacttagt acgcagaata
     2281 cgcactgcga actatttaat ttctactaat ggcgataaat tttcacaccc taacattgag
     2341 accttagcta gaatcattaa gttttctcct gaccgcgagg caaaaatact actaaaccac
     2401 gaaatccccc acctcacaaa ctccttcctg agcaacgcca acagttatga aggaaatatc
     2461 agcatccacc agggcataaa ggaatttgag ctttaataac attggaagtg gataacggat
     2521 ccgcgatcgc ggcgcgccac ctggtggccg gccggtacca cgcgtgcgcg ctgatccggc
     2581 tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc
     2641 ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaag gaggaactat
     2701 atccggatat ccacaggacg ggtgtggtcg ccatgatcgc gtagtcgata gtggctccaa
     2761 gtagcgaagc gagcaggact gggcggcggc caaagcggtc ggacagtgct ccgagaacgg
     2821 gtgcgcatag aaattgcatc aacgcatata gcgctagcag cacgccatag tgactggcga
     2881 tgctgtcgga atggacgata tcccgcaaga ggcccggcag taccggcata accaagccta
     2941 tgcctacagc atccagggtg acggtgccga ggatgacgat gagcgcattg ttagatttca
     3001 tacacggtgc ctgactgcgt tagcaattta actgtgataa actaccgcat taaagcttat
     3061 cgatgataag ctgtcaaaca tgagaattct tgaagacgaa agggcctcgt gatacgccta
     3121 tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg
     3181 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     3241 ctcatgagac aataaccctg ataaatgctt caataacatt gaaaaaggaa gagtatgagt
     3301 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     3361 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     3421 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     3481 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt
     3541 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     3601 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     3661 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     3721 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     3781 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgca
     3841 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     3901 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     3961 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     4021 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     4081 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     4141 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     4201 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     4261 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     4321 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     4381 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     4441 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     4501 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     4561 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     4621 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     4681 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     4741 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     4801 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     4861 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     4921 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     4981 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     5041 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     5101 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcaa tggtgcactc
     5161 tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg
     5221 tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     5281 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     5341 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc
     5401 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt
     5461 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc
     5521 ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc
     5581 gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact
     5641 ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac
     5701 tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca
     5761 gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag
     5821 actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt
     5881 gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag
     5941 gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg
     6001 ccaggaccca acgctgcccg agatgcgccg cgtgcggctg ctggagatgg cggacgcgat
     6061 ggatatgttc tgccaagggt tggtttgcgc attcacagtt ctccgcaaga attgattggc
     6121 tccaattctt ggagtggtga atccgttagc gaggtgccgc cggcttccat tcaggtcgag
     6181 gtggcccggc tccatgcacc gcgacgcaac gcggggaggc agacaaggta tagggcggcg
     6241 cctacaatcc atgccaaccc gttccatgtg ctcgccgagg cggcataaat cgccgtgacg
     6301 atcagcggtc caatgatcga agttaggctg gtaagagccg cgagcgatcc ttgaagctgt
     6361 ccctgatggt cgtcatctac ctgcctggac agcatggcct gcaacgcggg catcccgatg
     6421 ccgccggaag cgagaagaat cataatgggg aaggccatcc agcctcgcgt cgcgaacgcc
     6481 agcaagacgt agcccagcgc gtcggccgcc atgccggcga taatggcctg cttctcgccg
     6541 aaacgtttgg tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa gattccgaat
     6601 accgcaagcg acaggccgat catcgtcgcg ctccagcgaa agcggtcctc gccgaaaatg
     6661 acccagagcg ctgccggcac ctgtcctacg agttgcatga taaagaagac agtcataagt
     6721 gcggcgacga tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt gaaggctctc
     6781 aagggcatcg gtcgacgctc tcccttatgc gactcctgca ttaggaagca gcccagtagt
     6841 aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga gatggcgccc
     6901 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
     6961 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
     7021 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat cgagatctcg
     7081 atcccgcgaa a
//
