Molecular cloning of the gene encoding the mosaic neurotoxin, composed of parts of botulinum neurotoxin types C1 and D, and PCR detection of this gene from Clostridium botulinum type C organisms

Appl Environ Microbiol. 1996 Feb;62(2):662-7. doi: 10.1128/aem.62.2.662-667.1996.

Abstract

The DNA fragment common to the genes encoding botulinum neurotoxin types C1 (BN/C1) and D (BN/D) was amplified by PCR from the culture supernatant of Clostridium botulinum type C strain 6813 (C6813) that was treated with either DNase I or proteinase K but not from the supernatant that was treated with both DNase I and proteinase K, suggesting the neurotoxin gene is located on a certain bacteriophage DNA. Thus, to isolate the neurotoxin gene, we performed PCR with the culture supernatant of C6813 and seven primer pairs designed from the genes encoding BN/C1 and BN/D. The coding region in the connected sequence encodes a neurotoxin composed of 1,280 amino acids with a molecular weight of 147,817. The neurotoxin from C6813 has 95% amino acid identity to BN/C1, except for its C-terminal one-third, which is quite similar to the C-terminal one-third of BN/D (95% identity). When we performed PCRs with four primer pairs designed from the 5'-terminal two-thirds of the BN/C1 gene and two primers from the 3'-terminal one-third of the BN/D gene, DNA fragments of the expected sizes (0.5 to 1.3 kbp) could be amplified from C. botulinum type C strains 6812 and 6814. These results suggest that some strains of C. botulinum type C contain the gene encoding the mosaic neurotoxin composed of parts of BN/C1 and BN/D.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Botulinum Toxins / genetics*
  • Cloning, Molecular
  • Clostridium botulinum / classification
  • Clostridium botulinum / genetics*
  • DNA Primers / genetics
  • DNA, Bacterial / genetics
  • Genes, Bacterial*
  • Molecular Sequence Data
  • Mosaicism
  • Polymerase Chain Reaction
  • Sequence Homology, Amino Acid

Substances

  • DNA Primers
  • DNA, Bacterial
  • Botulinum Toxins

Associated data

  • GENBANK/D49440