We used a polymerase chain reaction (PCR) based cloning strategy to isolate cinnamomin genes from Phytophthora cinnamomi 8601, a pathogen responsible for cranberry root rot. Complete DNA sequence analysis of nine recombinant clones revealed two different classes of genes, each class consisting of genes with identical DNA sequences. Both classes of genes (Cin-1 and Cin-2) contained an open reading frame encoding a protein of 122 amino acid residues. The encoded proteins, named cinnamomin-1 and cinnamomin-2 (Cin-1 and Cin-2), were highly homologous to other proteins of the elicitin family and contained a 19 amino acid residue long signal peptide sequence. Both Cin-1 and Cin-2 proteins showed higher degree of sequence homology to the alpha-elicitins than beta-elicitins; moreover, a Val residue was found at position 13 of the putative mature Cin-1 and Cin-2 proteins. Because alpha-elicitins and beta-elicitins are known to contain a Val and a Lys residue, respectively, at this position, we concluded that both Cin-1 and Cin-2 genes from P. cinnamomi 8601 encode for alpha cinnamomins, Cin-1 and Cin-2.

Using primers based on the nucleotide sequence of a neurotoxin binding protein from Type E Clostridium botulinum cultures, an amplified DNA product was obtained through polymerase chain reaction. The 400 base pair amplified DNA fragment was detectable with as low as 0.1 pg template DNA from Type E C. botulinum, and its fidelity was confirmed by Southern blotting using a DNA probe designed to detect the expected amplified DNA fragment. On the other hand, no DNA amplification was observed with as high as 10 ng template DNA from related Types A and B C. botulinum or from C. tetani, indicating the specificity of the probe.

Botulinum and tetanus neurotoxins, produced by Clostridium botulinum and Clostridium tetani, respectively, are the most poisonous poisons known to mankind. Although botulinum and tetanus neurotoxins share several characteristics, such as similar mol. wts, similar macrostructure, virtually identical mode of action, and a strong amino acid sequence homology, the two neurotoxins differ in one very significant way; only botulinum neurotoxin is a food poison. Factors responsible for the food poisoning potential of botulinum neurotoxins seem to be a group of complexing proteins that are also produced by C. botulinum, and are known to associate with the neurotoxin. Translation products of nucleotide sequences upstream to the neurotoxin genes of serotypes A, B, C, D, E and F botulinum neurotoxin reveal the location of genes for one of the complexing proteins that could be transcribed as polycistronic mRNA to include neurotoxin sequences. No such protein seems to be present in C. tetani, suggesting that the lack of complexing proteins might be responsible for tetanus not being a food poison.