LOCUS       sequence-239496-        5559 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression vector for active 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 64.1
FEATURES             Location/Qualifiers
     source          1..5559
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    30..48
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             67..78
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      3069..3140
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3156..3183
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(3211..3228)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3345..3890)
                     /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(3425..3444)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(4004..4098)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(4119..4778)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4693..4712
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4779..4881)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             complement(4911..5537)
                     /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     5442..5461
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        join(5553..5559,1..49)
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
ORIGIN
        1 ctctatcgtt gatagagtta ttttaccact ccctatcagt gatagagaaa agaattcaaa
       61 agatctaaag aggagaaagg atctatggaa aagagaatca acaagatcag gaagaagctg
      121 agcgccgaca acgccaccaa gcctgtgtct aggagtggcc ccatgaaaac cctgctcgtc
      181 agagtgatga ccgacgacct gaagaagaga ctggaaaaga ggaggaagaa acccgaagtg
      241 atgccccaag tgatcagcaa caacgccgcc aacaacctga gaatgctgct ggacgactac
      301 accaagatga aggaagccat cctgcaggtc tactggcaag agttcaagga cgaccacgtg
      361 ggcctgatgt gcaagtttgc ccagcctgcc agcaagaaga tcgaccagaa caagctgaag
      421 cccgagatgg acgagaaggg caatctgacc acagccggct ttgcctgctc tcagtgtggc
      481 cagcctctgt tcgtgtacaa gctggaacag gtgtccgaga aaggcaaggc ctacaccaac
      541 tacttcggca gatgtaacgt ggccgagcac gagaagctga ttctgctggc ccagctgaaa
      601 cctgagaagg actctgatga ggccgtgacc tacagcctgg gcaagtttgg acagagagcc
      661 ctggacttct acagcatcca cgtgaccaaa gaaagcacac accccgtgaa gcccctggct
      721 cagatcgccg gcaatagata cgcctctgga cctgtgggca aagccctgtc cgatgcctgc
      781 atgggaacaa tcgccagctt cctgagcaag taccaggaca tcatcatcga gcaccagaag
      841 gtggtcaagg gcaaccagaa gagactggaa agcctgaggg agctggccgg caaagagaac
      901 ctggaatacc ccagcgtgac cctgcctcct cagcctcaca caaaagaagg cgtggacgcc
      961 tacaacgaag tgatcgccag agtgagaatg tgggtcaacc tgaacctgtg gcagaagctg
     1021 aaactgtcca gggacgacgc caagcctctg ctgagactga agggcttccc tagcttcccc
     1081 gtggtggaaa gaagggagaa tgaggtggac tggtggaaca ccatcaacga agtcaagaaa
     1141 ctgatcgatg ccaagagaga catgggcaga gtgttttgga gcggagtgac cgccgagaag
     1201 agaaacacca tcctggaagg ctacaactac ctgcctaacg agaacgacca caagaagaga
     1261 gagggcagcc tggaaaaccc taagaagccc gccaagagac agttcggcga cctgctgctg
     1321 tacctggaga agaagtatgc cggcgactgg ggcaaagtgt tcgatgaggc ctgggagaga
     1381 atcgacaaga agattgccgg cctgaccagc cacatcgaga gagaggaagc cagaaacgcc
     1441 gaggacgccc agtctaaagc cgtgctgaca gattggctga gagccaaggc cagcttcgtg
     1501 ctggaaaggc tgaaagaaat ggatgagaaa gagttctacg cctgcgagat ccagctgcag
     1561 aagtggtacg gggacctgag aggcaatccc tttgccgtgg aagccgagaa cagagtggtg
     1621 gacatcagcg gctttagcat cggctctgac ggccactcca tccagtacag gaacctgctg
     1681 gcctggaagt acctggaaaa tggcaagaga gagttctacc tgctgatgaa ctacggcaag
     1741 aagggcagaa tcagattcac cgacggcacc gacatcaaga aaagcggcaa gtggcagggc
     1801 ctgctgtatg gcggcggaaa ggccaaagtg atcgacctga ccttcgatcc cgacgacgag
     1861 cagctgatca ttctgcctct ggccttcggc accagacagg gcagagagtt catctggaac
     1921 gatctgctgt ccctggaaac cggcctgatc aagctggcca acggcagggt gatcgagaaa
     1981 accatctaca acaagaaaat cggcagagat gagcccgcac tgtttgtggc cctgaccttt
     2041 gagaggagag aagtggtcga tcccagcaac atcaagcccg tgaacctgat cggagtggac
     2101 agaggcgaga atatccctgc cgtgattgcc ctgacagacc ctgagggatg tcccctgcct
     2161 gagtttaagg atagctctgg cggccctacc gacatcctga gaatcggcga gggctacaaa
     2221 gagaagcaga gagccatcca ggccgccaaa gaggtggaac agagaagagc cggcggatac
     2281 agcagaaagt tcgccagcaa gtccagaaac ctggccgatg acatggtcag gaacagcgcc
     2341 agggacctgt tctaccacgc cgtgacacat gatgccgtgc tggtgttcga gaacctgtca
     2401 agaggcttcg gcagacaggg aaaaagaacc ttcatgaccg agagacagta cacgaagatg
     2461 gaagattggc tcaccgccaa actggcctac gagggcctga caagcaagac ctacctgtcc
     2521 aagacactgg cccagtacac ctccaagacc tgctccaatt gcggcttcac catcaccacc
     2581 gccgactacg atggaatgct cgtgagactg aagaaaacct ccgacggctg ggccaccaca
     2641 ctgaacaaca aagagctgaa agccgagggc cagatcacct actacaacag atacaagagg
     2701 cagaccgtcg agaaagagct gtccgccgaa ctcgacagac tgagcgagga atccggcaac
     2761 aacgacatca gcaagtggac caagggacgc agagatgagg ccctgttcct gctgaagaag
     2821 aggttcagcc acagacctgt gcaagagcag tttgtgtgcc tggattgcgg ccacgaagtg
     2881 cacgcagatg aacaggccgc tctgaatatc gccagaagct ggctgttcct gaacagcaac
     2941 agcaccgagt tcaagagcta caagagcggc aagcagcctt tcgttggagc ctggcaggcc
     3001 ttctacaaga gaaggctcaa agaagtgtgg aagcccaacg cctgactcga gtaaggatct
     3061 ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt
     3121 gtttgtcggt gaacgctctc tactagagtc acactggctc accttcgggt gggcctttct
     3181 gcgtttatac ctagggatat attccgcttc ctcgctcact gactcgctac gctcggtcgt
     3241 tcgactgcgg cgagcggaaa tggcttacga acggggcgga gatttcctgg aagatgccag
     3301 gaagatactt aacagggaag tgagagggcc gcggcaaagc cgtttttcca taggctccgc
     3361 ccccctgaca agcatcacga aatctgacgc tcaaatcagt ggtggcgaaa cccgacagga
     3421 ctataaagat accaggcgtt tccccctggc ggctccctcg tgcgctctcc tgttcctgcc
     3481 tttcggttta ccggtgtcat tccgctgtta tggccgcgtt tgtctcattc cacgcctgac
     3541 actcagttcc gggtaggcag ttcgctccaa gctggactgt atgcacgaac cccccgttca
     3601 gtccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg aaagacatgc
     3661 aaaagcacca ctggcagcag ccactggtaa ttgatttaga ggagttagtc ttgaagtcat
     3721 gcgccggtta aggctaaact gaaaggacaa gttttggtga ctgcgctcct ccaagccagt
     3781 tacctcggtt caaagagttg gtagctcaga gaaccttcga aaaaccgccc tgcaaggcgg
     3841 ttttttcgtt ttcagagcaa gagattacgc gcagaccaaa acgatctcaa gaagatcatc
     3901 ttattaatca gataaaatat ttctagattt cagtgcaatt tatctcttca aatgtagcac
     3961 ctgaagtcag ccccatacga tataagttgt tactagtgct tggattctca ccaataaaaa
     4021 acgcccggcg gcaaccgagc gttctgaaca aatccagatg gagttctgag gtcattactg
     4081 gatctatcaa caggagtcca agcgagctcg atatcaaatt acgccccgcc ctgccactca
     4141 tcgcagtact gttgtaattc attaagcatt ctgccgacat ggaagccatc acaaacggca
     4201 tgatgaacct gaatcgccag cggcatcagc accttgtcgc cttgcgtata atatttgccc
     4261 atggtgaaaa cgggggcgaa gaagttgtcc atattggcca cgtttaaatc aaaactggtg
     4321 aaactcaccc agggattggc tgacacgaaa aacatattct caataaaccc tttagggaaa
     4381 taggccaggt tttcaccgta acacgccaca tcttgcgaat atatgtgtag aaactgccgg
     4441 aaatcgtcgt ggtattcact ccagagcgat gaaaacgttt cagtttgctc atggaaaacg
     4501 gtgtaacaag ggtgaacact atcccatatc accagctcac cgtctttcat tgccatacga
     4561 aattccggat gagcattcat caggcgggca agaatgtgaa taaaggccgg ataaaacttg
     4621 tgcttatttt tctttacggt ctttaaaaag gccgtaatat ccagctgaac ggtctggtta
     4681 taggtacatt gagcaactga ctgaaatgcc tcaaaatgtt ctttacgatg ccattgggat
     4741 atatcaacgg tggtatatcc agtgattttt ttctccattt tagcttcctt agctcctgaa
     4801 aatctcgata actcaaaaaa tacgcccggt agtgatctta tttcattatg gtgaaagttg
     4861 gaacctctta cgtgccgatc aacgtttcat tttcgccaga tatcgacgtc ttaagaccca
     4921 ctttcacatt taagttgttt ttctaatccg catatgatca attcaaggcc gaataagaag
     4981 gctggctctg caccttggtg atcaaataat tcgatagctt gtcgtaataa tggcggcata
     5041 ctatcagtag taggtgtttc cctttcttct ttagcgactt gatgctcttg atcttccaat
     5101 acgcaaccta aagtaaaatg ccccacagcg ctgagtgcat ataatgcatt ctctagtgaa
     5161 aaaccttgtt ggcataaaaa ggctaattga ttttcgagag tttcatactg tttttctgta
     5221 ggccgtgtac ctaaatgtac ttttgctcca tcgcgatgac ttagtaaagc acatctaaaa
     5281 cttttagcgt tattacgtaa aaaatcttgc cagctttccc cttctaaagg gcaaaagtga
     5341 gtatggtgcc tatctaacat ctcaatggct aaggcgtcga gcaaagcccg cttatttttt
     5401 acatgccaat acaatgtagg ctgctctaca cctagcttct gggcgagttt acgggttgtt
     5461 aaaccttcga ttccgacctc attaagcagc tctaatgcgc tgttaatcac tttactttta
     5521 tctaatctag acatcattaa ttcctaattt ttgttgaca
//
