THE CHROMOSOME ORGANIZATION OF GENES AND SOME TYPES OF EXTRAGENIC DNA IN ALLIUM

Abstract

We have studied the physical organization of genes and non-coding DNA in two Allium species, A. cepa and A. fistulosum. Expressed sequence tags (ESTs) clones and polymerase chain reaction (PCR) products of gene fragments obtained with primers designed for the gene sequences available in the public GenBank were physically mapped onto chromosomes using a Tyr-FISH technique. The allinase and lacrymatory factor synthase (LFS) genes that encoded enzymes operating in one metabolic way were cloned sequences and physically mapped onto A. cepa and A. fistulosum chromosomes. The alliinase gene probe (1100 bp) hybridized to the distal region of the long arm of chromosome 4 of A. cepa. The LFS gene probe (550 bp) hybridized to the proximal region of the long arm of chromosome 5 in both species. Inter-simple sequence repeats (ISSRs) that are located between SSR loci were used for chromosomal location of microsatellites in A. fistulosum using common FISH. The chromosomal organization of the Tyl-copia group retrotransposons were investigated in A. fistulosum. Dispersed hybridization of the probe along the chromosome arms apart from telomeric ends was detected. Chromosomal distribution of DNA methylation pattern in A. fistulosum L. was studied using a specific antibodies against 5-methylcytosine (anti-5mC). Highly methylated distal regions in all chromosomes were found. Differences in the methylation level between corresponding regions of homologue chromosomes were shown. We describe recent progress in exploiting the ultrasensitive Tyr-FISH technique for development of visual gene maps for chromosomes of A. cepa and A. fistulosum. The results on the chromosomal location of individual genetic loci aided in assembling physical and genetic maps. We related the physical organization of expressed genes to the contrasting patterns of chiasma distribution and to the organization of repetitive DNA family and highly methylated DNA in these two Allium species. INTRODUCTION A half century ago we learned that higher eukaryotic organisms possess much more DNA in their genome than they likely need for genetic information (MacLean, 1973). In onion, the difference between gene fraction (3-5% of the genome) and extragenic DNA fraction (93-97%) is extremely high (Flavell et al., 1974; Stack and Comings, 1979). Onion has one of the largest genomes among monocots (16 415 Mbp per 1C nucleus). The extraordinary huge genome of onion delays the construction of genomic resources for this economically important crop. Onion is used in every home, daily. Onion is the second most valued vegetable crop (FAO, 2010). In Russia in 2010, onion was cultivated on 88,000 ha with total onion production mounted 1,536,300 tons (FAO, 2010). The best way to get a comprehensive insight into chromosome organization of genes and extragenic DNA is the genome sequencing of species. With the development of second generation sequencing techniques that are capable of sequencing thousands of a Corresponding author. Proc. 6 IS on Edible Alliaceae