LOCUS       sequence-500767-        7717 bp    DNA     circular SYN 23-FEB-2026
DEFINITION  Hachiman from P. fluorescens under pTet control with HamM mutated 
            (I118K).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7717)
  AUTHORS   Doudna J et al.
  TITLE     Recurrent acquisition of nuclease-protease pairs in antiviral 
            immunity
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7717)
  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            "Unpublished"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: pTet_PflHamM(I118K)AB
FEATURES             Location/Qualifiers
     source          1..7717
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             133..162
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             178..195
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(251..268)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      421..467
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(499..1125)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     1030..1049
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        1141..1196
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    1177..1195
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      complement(6001..6035)
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     CDS             complement(6106..6765)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     6680..6699
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     complement(7004..7021)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7138..7683)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(7218..7237)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agatttcagt gcaatttatc tcttcaaatg tagcacctga agtcagcccc atacgatata
       61 agttgtcacc tgcccgaacc agagacttat cctgaactgg ccgataattg cagacgaacg
      121 atgcttgggc ccgaacaaaa actcatctca gaagaggatc tgaatagcgc cgtcgaccat
      181 catcatcatc atcattgagt ttaaacgctc tccagcttgg gtgttttggc ggatgagaga
      241 agattttcag cctgatacag attaaatcag aacgcagaag cggtctgata aaacagaatt
      301 tgcctggcgg cagtagcgcg gtggtcccac ctgaccccat gccgaactca gaagtgaaac
      361 gccgtagcgc cgatggtagt gtggggtctg cccatgcgag agtaggcaac tgccaggcat
      421 caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg
      481 gtgaacttca gcgacgtctt aagacccact ttcacattta agttgttttt ctaatccgca
      541 tatgatcaat tcaaggccga ataagaaggc tggctctgca ccttggtgat caaataattc
      601 gatagcttgt cgtaataatg gcggcatact atcagtagta ggtgtttccc tttcttcttt
      661 agcgacttga tgctcttgat cttccaatac gcaacctaaa gtaaaatgcc ccacagcgct
      721 gagtgcatat aatgcattct ctagtgaaaa accttgttgg cataaaaagg ctaattgatt
      781 ttcgagagtt tcatactgtt tttctgtagg ccgtgtacct aaatgtactt ttgctccatc
      841 gcgatgactt agtaaagcac atctaaaact tttagcgtta ttacgtaaaa aatcttgcca
      901 gctttcccct tctaaagggc aaaagtgagt atggtgccta tctaacatct caatggctaa
      961 ggcgtcgagc aaagcccgct tattttttac atgccaatac aatgtaggct gctctacacc
     1021 tagcttctgg gcgagtttac gggttgttaa accttcgatt ccgacctcat taagcagctc
     1081 taatgcgctg ttaatcactt tacttttatc taatctagac atcattaatt cctaattttt
     1141 gttgacactc tatcgttgat agagttattt taccactccc tatcagtgat agagaaaaga
     1201 attcaaaaga tcttttaaga aggagatata ctgatgaaaa tttccatgca cgatgccgca
     1261 tacggtgatt caattctcat cagcggcgcc tcgaccaaaa ttttagtgga tggtggaact
     1321 gcgaagtcct tcgctgactg gtcgccttcg ttagagggtg tcgatacgct agatgccctg
     1381 atagttactc atattgataa cgaccatatt ggcggagcaa tcaagctgct cacatcaaag
     1441 cacgcaacta aagtaaaaga agtttatttc aatggcatta accaaatcct agatgtggag
     1501 gattgtgata ttggtgatcg tagtgacgat ctagaattat ccgcaatcat agcaaacacc
     1561 acgtctagag acgaagacca gttgaaaggg tattctgaag gcacatcgtt atccttaata
     1621 ttacagcagc aaaatatccc cttaaataaa gcaaccgaag gcaggcaaat ccacagaacg
     1681 agccttaata aaaacataat aataggagaa tttgaaattg aactgatagg cccaagcaga
     1741 aagagcttgt taaaaatcca acaaaaatgg cttgaactat tagaaagtaa agaaatcaaa
     1801 agacgcctta tcagcccaag atatagcgtt gcattcgaaa gctttatcag ctcgattgat
     1861 acctcagtaa agcatgatga ggaaatttca agtctaagtg aaaacataga gcatttggca
     1921 aatcaaccat ttactaatga cacatcaccc ccaaacaaaa gcagcatctc tttccttcta
     1981 aaaactgaaa acaaaagcat tttaatgctc ggagattccg acgcagagac tatagaggca
     2041 tggttgaact cgaagtctgc agatagcatc acagttgatg cggtaaagct accccatcac
     2101 ggaagccgaa ataactttag tgctaactta gtacgcagaa tacgcactgc gaactattta
     2161 atttctacta atggcgataa attttcacac cctaacattg agaccttagc tagaatcatt
     2221 aagttttctc ctgaccgcga ggcaaaaata ctactaaacc acgaaatccc ccacctcaca
     2281 aactccttcc tgagcaacgc caacagttat gaaggaaata tcagcatcca ccagggcata
     2341 aaggaatttg agctttgagt aatgcactga ctgacgagta caaactcgat aacgtcttta
     2401 tcctagaagc ctatgacgac ctaggtgaac ccatttcaac aggcagttgc ttttcggtgg
     2461 gtgcgtctat tttttttaca tcttggcacg tcgtcaaaaa tgcctacacg cttcggattt
     2521 tccaaagtag agatgattat tttcgtcaag agttccatgc cgctaaatgt gttttttacg
     2581 ataaagaaac tgacttcgca atactgtcat cagacagcct tacgatcaaa tcaaaaattg
     2641 aaataggaat atacaaagct gagctgaatg atgaggtaaa agctataggg tacgcggccg
     2701 aaaaaagcag attgcatttc cctataacta gtaaaatatc aaacgtcttt gatttctcta
     2761 caacatataa atatgacttt gagtttggaa agccggctga gctagacaaa gcaaatggct
     2821 tatccggggg gccagtattc tttaaaggct acgctattgg cttgcttgca ctgcaaactg
     2881 tgggatcggc attgcaagca gtatctttcc atacaatatt tgaacggcca tccgctcttg
     2941 aagccattga aagccaaggg ttggttataa ccggtggtga cgaaatatcc tatgataggc
     3001 cagaccatcc tgaaagccca tttgttgtct caatagactg tgccgcttcg gatccaaaca
     3061 taaagggact agatatagga tttgatcttg gcgaatggcg cagtcagaac ctgattgatc
     3121 aaacgctcaa ctggatcgaa gactacgcgc tgaccccgaa gcaaaaaaaa gctttggggc
     3181 aacacaactt caaaagaatt caaagtgcaa aaagaaactt ccagattggc aacccagcag
     3241 caatggccga tttattctta cacatcgcga ttcgccaaaa ttataaaacc atccctatta
     3301 tttcaaatgt aataacggta gatggcgagt ctgtgttttc ctgctcgcat atcgttattt
     3361 atagaggcaa agttgagcta tggcttggaa tctctgctat ggaggtttcc atggagcaaa
     3421 cagtaaagga ggctattcac agagttgact ccatcatacg ttccggagga atccaagacc
     3481 ggttggttgt tatattatca gaactagacc ctacttggcc gtatataaaa aagctggaaa
     3541 aattatcaaa ctccaacctc aaattaactg acaaaattga taagataatt atacctatgt
     3601 ttttatcaag caatgcagac ttgataaata attatgaccc aagcaatttt tcaaactcat
     3661 ttcaaaaaga tatagatgag tgcagggaaa gggttgctgg cgcctacaat gctcgaatcg
     3721 ttcccttgat caacctccgc atctttctat ttccggtcga caacatacac gagttacata
     3781 cgaaatttgt taaggagttc gaattttgaa tacggaaatt tataacaacc ttcgtaagca
     3841 aatcgacaac tctgctttca ccggatggca aaggttattc gttgaactta taaatctata
     3901 caacttacat gatgacaatc aattcttgat tcaggacctg acgctgagat gcctagagag
     3961 gcgtggggaa ttcaaacagc ttcatggtct gttgaacata ttgatttccg agctaggttt
     4021 gttcccgtac ttagatccag atctgatgac gtcgaaagac agcataagga tgaatatttt
     4081 taagtcgcct atagctggcg atataacatt ccacctaaaa caggctgaga tttattacag
     4141 gattgtaaat ggagataaca tcgtactcag tgcacctaca agcttcggaa aaagcttagt
     4201 tatagatgct ctcatcagct ctgaagaata cgacaacatt gtggtagtag tcccaaccat
     4261 agctttgatg gatgagttac gaaagaaatt tagcaaatat aaagaaaact acaagataat
     4321 cactcagtcc tctcaagagc cagcgacaaa aaatctatat atctatactc aagaaagggt
     4381 cttggaatgc aagaaccttg aacgagtaga cttctttatt atcgatgagt tttacaaatt
     4441 agcacctctc aatacatctg actatcgaag cgacagacta aatctggcat ttcacaaact
     4501 ttttaaaatg tgcagtcgct tttacatgct cggccccaac attaatggcc tggccgaagg
     4561 catagaagac aaactaaatt gcaattttgt aaaatatgac aattacaaaa ctgtagtagc
     4621 taacgaatat cgctacatta tccgtgaaga tgggaatgat aaagaaaaag acattgatcg
     4681 cgacgcgcac ttagtcaata ttttgcaaaa aatcggtcaa aacgaacaga ctttaattta
     4741 ctgcaagact ccaaaacgag cagcctcgct agtaaaaaag atcatgaaac tcagagagaa
     4801 ccaagagata agaaaaggag agttttctga ctggctgaga gagacatatc acgatgactg
     4861 gtcgatggca gacgcagtgg aacgcgggat tgcctgtcac catggcagac ttcctcgtgt
     4921 cgtctccggg tatgtggtgg aagcgttcaa tgaaaagctc ataaatattt tggtttgcac
     4981 ctcaactttg attgagggtg ttaacacaaa tgctagaagc gtcataattt acgatgactg
     5041 tattcgaggg accacccaac tggattcgtt cacttttaac aatatctctg gacgtagtgg
     5101 gcgtatgttt gaacactttg ttggagatat acatgttatt ggacgaaaac catcagcaga
     5161 gcttccttgg atcgacgtcc caatagtgac tcaagatgac tccgcctcta gacccatgct
     5221 gctaagcata atcgacgagc taagcaccga acaaagggct agtttatcga gatatacaac
     5281 tcaacaatat gtacctatca ctttacttaa taagcatctc ggtgttcctc cggaaaggat
     5341 aataaaattc gcagaagcct taatggagaa atgtggcgcc tggaactcaa ttatggcatg
     5401 gacgggaaaa ttcccaacta gcgagcaact aagacaccta tgcaacatca tgttcgatca
     5461 cttcgcaata tcacgaatgg gcggaggatg cgtcaaatca aaaaagcagc tgttttatag
     5521 agtgctcaac attgtaaata gaaaagaaga caaagatcta attcaaagcg aatatgattt
     5581 ttggtcccca cgaaacgcgg aatatgagat tgatgacgca gtgcaaagcg ttttcgattt
     5641 caaaagaaac ttaattggct acaacctccc taagattatt ttggcgataa acgacgtaca
     5701 gcgcttaata ttttctcgat tcaattaccg ttgtggcgac tacgcagctt ttgcccactc
     5761 tctagaagca atgtttttac caccttcaat tgtggctctt gaagagtttg gaataccaac
     5821 tcgtgtagca caaaaaatat accaaattag caacgtcggc gaggacgcca ccgtagattt
     5881 aatcctagag caggtaagat cgcacccggg tctctacata tcgaaatttg atgattttga
     5941 aaaaactctc ctgaaccgag caattcaata tctttaaagg aggactttgg caggtgacca
     6001 ataaaaaacg cccggcggca accgagcgtt ctgaacaaat ccagatggag ttctgaggtc
     6061 attactggat ctatcaacag gagtccaagc gagctcgata tcaaattacg ccccgccctg
     6121 ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga agccatcaca
     6181 aacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt gcgtataata
     6241 tttgcccatg gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt ttaaatcaaa
     6301 actggtgaaa ctcacccagg gattggctga aacgaaaaac atattctcaa taaacccttt
     6361 agggaaatag gccaggtttt caccgtaaca cgccacatct tgcgaatata tgtgtagaaa
     6421 ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag tttgctcatg
     6481 gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt ctttcattgc
     6541 catacgaaat tccggatgag cattcatcag gcgggcaaga atgtgaataa aggccggata
     6601 aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca gctgaacggt
     6661 ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt tacgatgcca
     6721 ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag cttccttagc
     6781 tcctgaaaat ctcgataact caaaaaatac gcccggtagt gatcttattt cattatggtg
     6841 aaagttggaa cctcttacgt gcccgatcaa gcgctagcgg agtgtatact ggcttactat
     6901 gttggcactg atgagggtgt cagtgaagtg cttcatgtgg caggagaaaa aaggctgcac
     6961 cggtgcgtca gcagaatatg tgatacagga tatattccgc ttcctcgctc actgactcgc
     7021 tacgctcggt cgttcgactg cggcgagcgg aaatggctta cgaacggggc ggagatttcc
     7081 tggaagatgc caggaagata cttaacaggg aagtgagagg gccgcggcaa agccgttttt
     7141 ccataggctc cgcccccctg acaagcatca cgaaatctga cgctcaaatc agtggtggcg
     7201 aaacccgaca ggactataaa gataccaggc gtttccccct ggcggctccc tcgtgcgctc
     7261 tcctgttcct gcctttcggt ttaccggtgt cattccgctg ttatggccgc gtttgtctca
     7321 ttccacgcct gacactcagt tccgggtagg cagttcgctc caagctggac tgtatgcacg
     7381 aaccccccgt tcagtccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     7441 cggaaagaca tgcaaaagca ccactggcag cagccactgg taattgattt agaggagtta
     7501 gtcttgaagt catgcgccgg ttaaggctaa actgaaagga caagttttgg tgactgcgct
     7561 cctccaagcc agttacctcg gttcaaagag ttggtagctc agagaacctt cgaaaaaccg
     7621 ccctgcaagg cggttttttc gttttcagag caagagatta cgcgcagacc aaaacgatct
     7681 caagaagatc atcttattaa tcagataaaa tatttct
//
