Southern blot and sequence analysis suggest multiple copies of the alliinase gene in the garlic (Allium sativum l.) genome

CAVAGNARO, P. F.; MASUELLI, R. W.; SIMON, P. W.

Conferencia; Plant & Animal Genomes XI Conference; 2003

Alliinase is an important enzyme found in Alliaceae species. Alliinase activity in garlic is involved in the production of organosulfur compounds responsable for todor, pungency, health benefits, and resistance to several plant pathogens.Van Damme et al. found sequence differences among garlic bulb cDNA-alliinase clones, suggesting that this enzyme is encoded by a family of genes. To investigate the number of copies of alliinase-encoding genes per garlic genome we performed Southern Blot analysis, using a P32 labelled allinase cDNA probe in 22 garlic genotypes. Four to eight bands per garlic clone were observed, suggesting the existence of, at least, the same number of copies per genome. Specific primers were designed and used to amplify an intron-bearing fragment of the gene, in garlic clone DDR7040. PCR products were cloned and 200 recombinant colonies were sequenced. More than 25 sequence variants were identified, by means of SNPs and small deletions. It is posible that, due to errors during PCR amplification, we could be overstimating the amount of gene variants. The sequence of a few complete alliinase genes showed an intronless variant in contrast to the 4 introns usually observed. Our results suggest multiple copies comprising the alliinase gene family. The posibility that many of them could be functional, agrees with the high content of this enzime in garlic bulbs. Presumably, the amplification of this gene in the garlic genome, could have had an important role in the adaptiveness and evolution of this species.