LOCUS       sequence-235007-        5559 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression vector for catalytically inactivated 
            deltaproteobacteria CasX.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5559)
  AUTHORS   Liu JJ, Orlova N, Oakes BL, Ma E, Spinner HB, Baney KLM, Chuck J, 
            Tan D, Knott GJ, Harrington LB, Al-Shayeb B, Wagner A, Brotzmann J, 
            Staahl BT, Taylor KL, Desmarais J, Nogales E, Doudna JA
  TITLE     CasX enzymes comprise a distinct family of RNA-guided genome 
            editors.
  JOURNAL   Nature. 2019 Feb 4. pii: 10.1038/s41586-019-0908-x. doi: 
            10.1038/s41586-019-0908-x.
  PUBMED    30718774
REFERENCE   2  (bases 1 to 5559)
  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: "Nature. 
            2019 Feb 4. pii: 10.1038/s41586-019-0908-x. doi: 
            10.1038/s41586-019-0908-x."
COMMENT     Sequence Label: pBLO ddd64.1
FEATURES             Location/Qualifiers
     source          1..5559
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(7..633)
                     /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     538..557
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        649..704
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    685..703
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             722..733
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      3724..3795
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3811..3838
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(3866..3883)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4000..4545)
                     /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(4080..4099)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(4659..4753)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(4774..5433)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5348..5367
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(5434..5536)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
ORIGIN
        1 gacgtcttaa gacccacttt cacatttaag ttgtttttct aatccgcata tgatcaattc
       61 aaggccgaat aagaaggctg gctctgcacc ttggtgatca aataattcga tagcttgtcg
      121 taataatggc ggcatactat cagtagtagg tgtttccctt tcttctttag cgacttgatg
      181 ctcttgatct tccaatacgc aacctaaagt aaaatgcccc acagcgctga gtgcatataa
      241 tgcattctct agtgaaaaac cttgttggca taaaaaggct aattgatttt cgagagtttc
      301 atactgtttt tctgtaggcc gtgtacctaa atgtactttt gctccatcgc gatgacttag
      361 taaagcacat ctaaaacttt tagcgttatt acgtaaaaaa tcttgccagc tttccccttc
      421 taaagggcaa aagtgagtat ggtgcctatc taacatctca atggctaagg cgtcgagcaa
      481 agcccgctta ttttttacat gccaatacaa tgtaggctgc tctacaccta gcttctgggc
      541 gagtttacgg gttgttaaac cttcgattcc gacctcatta agcagctcta atgcgctgtt
      601 aatcacttta cttttatcta atctagacat cattaattcc taatttttgt tgacactcta
      661 tcgttgatag agttatttta ccactcccta tcagtgatag agaaaagaat tcaaaggatc
      721 caaagaggag aaaggatcta tggaaaagag aatcaacaag atcaggaaga agctgagcgc
      781 cgacaacgcc accaagcctg tgtctaggag tggccccatg aaaaccctgc tcgtcagagt
      841 gatgaccgac gacctgaaga agagactgga aaagaggagg aagaaacccg aagtgatgcc
      901 ccaagtgatc agcaacaacg ccgccaacaa cctgagaatg ctgctggacg actacaccaa
      961 gatgaaggaa gccatcctgc aggtctactg gcaagagttc aaggacgacc acgtgggcct
     1021 gatgtgcaag tttgcccagc ctgccagcaa gaagatcgac cagaacaagc tgaagcccga
     1081 gatggacgag aagggcaatc tgaccacagc cggctttgcc tgctctcagt gtggccagcc
     1141 tctgttcgtg tacaagctgg aacaggtgtc cgagaaaggc aaggcctaca ccaactactt
     1201 cggcagatgt aacgtggccg agcacgagaa gctgattctg ctggcccagc tgaaacctga
     1261 gaaggactct gatgaggccg tgacctacag cctgggcaag tttggacaga gagccctgga
     1321 cttctacagc atccacgtga ccaaagaaag cacacacccc gtgaagcccc tggctcagat
     1381 cgccggcaat agatacgcct ctggacctgt gggcaaagcc ctgtccgatg cctgcatggg
     1441 aacaatcgcc agcttcctga gcaagtacca ggacatcatc atcgagcacc agaaggtggt
     1501 caagggcaac cagaagagac tggaaagcct gagggagctg gccggcaaag agaacctgga
     1561 ataccccagc gtgaccctgc ctcctcagcc tcacacaaaa gaaggcgtgg acgcctacaa
     1621 cgaagtgatc gccagagtga gaatgtgggt caacctgaac ctgtggcaga agctgaaact
     1681 gtccagggac gacgccaagc ctctgctgag actgaagggc ttccctagct tccccgtggt
     1741 ggaaagaagg gagaatgagg tggactggtg gaacaccatc aacgaagtca agaaactgat
     1801 cgatgccaag agagacatgg gcagagtgtt ttggagcgga gtgaccgccg agaagagaaa
     1861 caccatcctg gaaggctaca actacctgcc taacgagaac gaccacaaga agagagaggg
     1921 cagcctggaa aaccctaaga agcccgccaa gagacagttc ggcgacctgc tgctgtacct
     1981 ggagaagaag tatgccggcg actggggcaa agtgttcgat gaggcctggg agagaatcga
     2041 caagaagatt gccggcctga ccagccacat cgagagagag gaagccagaa acgccgagga
     2101 cgcccagtct aaagccgtgc tgacagattg gctgagagcc aaggccagct tcgtgctgga
     2161 aaggctgaaa gaaatggatg agaaagagtt ctacgcctgc gagatccagc tgcagaagtg
     2221 gtacggggac ctgagaggca atccctttgc cgtggaagcc gagaacagag tggtggacat
     2281 cagcggcttt agcatcggct ctgacggcca ctccatccag tacaggaacc tgctggcctg
     2341 gaagtacctg gaaaatggca agagagagtt ctacctgctg atgaactacg gcaagaaggg
     2401 cagaatcaga ttcaccgacg gcaccgacat caagaaaagc ggcaagtggc agggcctgct
     2461 gtatggcggc ggaaaggcca aagtgatcga cctgaccttc gatcccgacg acgagcagct
     2521 gatcattctg cctctggcct tcggcaccag acagggcaga gagttcatct ggaacgatct
     2581 gctgtccctg gaaaccggcc tgatcaagct ggccaacggc agggtgatcg agaaaaccat
     2641 ctacaacaag aaaatcggca gagatgagcc cgcactgttt gtggccctga cctttgagag
     2701 gagagaagtg gtcgatccca gcaacatcaa gcccgtgaac ctgatcggag tggccagagg
     2761 cgagaatatc cctgccgtga ttgccctgac agaccctgag ggatgtcccc tgcctgagtt
     2821 taaggatagc tctggcggcc ctaccgacat cctgagaatc ggcgagggct acaaagagaa
     2881 gcagagagcc atccaggccg ccaaagaggt ggaacagaga agagccggcg gatacagcag
     2941 aaagttcgcc agcaagtcca gaaacctggc cgatgacatg gtcaggaaca gcgccaggga
     3001 cctgttctac cacgccgtga cacatgatgc cgtgctggtg ttcgcgaacc tgtcaagagg
     3061 cttcggcaga cagggaaaaa gaaccttcat gaccgagaga cagtacacga agatggaaga
     3121 ttggctcacc gccaaactgg cctacgaggg cctgacaagc aagacctacc tgtccaagac
     3181 actggcccag tacacctcca agacctgctc caattgcggc ttcaccatca ccaccgccga
     3241 ctacgatgga atgctcgtga gactgaagaa aacctccgac ggctgggcca ccacactgaa
     3301 caacaaagag ctgaaagccg agggccagat cacctactac aacagataca agaggcagac
     3361 cgtcgagaaa gagctgtccg ccgaactcga cagactgagc gaggaatccg gcaacaacga
     3421 catcagcaag tggaccaagg gacgcagaga tgaggccctg ttcctgctga agaagaggtt
     3481 cagccacaga cctgtgcaag agcagtttgt gtgcctggat tgcggccacg aagtgcacgc
     3541 agctgaacag gccgctctga atatcgccag aagctggctg ttcctgaaca gcaacagcac
     3601 cgagttcaag agctacaaga gcggcaagca gcctttcgtt ggagcctggc aggccttcta
     3661 caagagaagg ctcaaagaag tgtggaagcc caacgcctga ctcgagtaag gatctccagg
     3721 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
     3781 tcggtgaacg ctctctacta gagtcacact ggctcacctt cgggtgggcc tttctgcgtt
     3841 tatacctagg gatatattcc gcttcctcgc tcactgactc gctacgctcg gtcgttcgac
     3901 tgcggcgagc ggaaatggct tacgaacggg gcggagattt cctggaagat gccaggaaga
     3961 tacttaacag ggaagtgaga gggccgcggc aaagccgttt ttccataggc tccgcccccc
     4021 tgacaagcat cacgaaatct gacgctcaaa tcagtggtgg cgaaacccga caggactata
     4081 aagataccag gcgtttcccc ctggcggctc cctcgtgcgc tctcctgttc ctgcctttcg
     4141 gtttaccggt gtcattccgc tgttatggcc gcgtttgtct cattccacgc ctgacactca
     4201 gttccgggta ggcagttcgc tccaagctgg actgtatgca cgaacccccc gttcagtccg
     4261 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggaaaga catgcaaaag
     4321 caccactggc agcagccact ggtaattgat ttagaggagt tagtcttgaa gtcatgcgcc
     4381 ggttaaggct aaactgaaag gacaagtttt ggtgactgcg ctcctccaag ccagttacct
     4441 cggttcaaag agttggtagc tcagagaacc ttcgaaaaac cgccctgcaa ggcggttttt
     4501 tcgttttcag agcaagagat tacgcgcaga ccaaaacgat ctcaagaaga tcatcttatt
     4561 aatcagataa aatatttcta gatttcagtg caatttatct cttcaaatgt agcacctgaa
     4621 gtcagcccca tacgatataa gttgttacta gtgcttggat tctcaccaat aaaaaacgcc
     4681 cggcggcaac cgagcgttct gaacaaatcc agatggagtt ctgaggtcat tactggatct
     4741 atcaacagga gtccaagcga gctcgatatc aaattacgcc ccgccctgcc actcatcgca
     4801 gtactgttgt aattcattaa gcattctgcc gacatggaag ccatcacaaa cggcatgatg
     4861 aacctgaatc gccagcggca tcagcacctt gtcgccttgc gtataatatt tgcccatggt
     4921 gaaaacgggg gcgaagaagt tgtccatatt ggccacgttt aaatcaaaac tggtgaaact
     4981 cacccaggga ttggctgaca cgaaaaacat attctcaata aaccctttag ggaaataggc
     5041 caggttttca ccgtaacacg ccacatcttg cgaatatatg tgtagaaact gccggaaatc
     5101 gtcgtggtat tcactccaga gcgatgaaaa cgtttcagtt tgctcatgga aaacggtgta
     5161 acaagggtga acactatccc atatcaccag ctcaccgtct ttcattgcca tacgaaattc
     5221 cggatgagca ttcatcaggc gggcaagaat gtgaataaag gccggataaa acttgtgctt
     5281 atttttcttt acggtcttta aaaaggccgt aatatccagc tgaacggtct ggttataggt
     5341 acattgagca actgactgaa atgcctcaaa atgttcttta cgatgccatt gggatatatc
     5401 aacggtggta tatccagtga tttttttctc cattttagct tccttagctc ctgaaaatct
     5461 cgataactca aaaaatacgc ccggtagtga tcttatttca ttatggtgaa agttggaacc
     5521 tcttacgtgc cgatcaacgt ttcattttcg ccagatatc
//
