Whole genome profiling physical map and ancestral annotation of tobacco Hicks Broadleaf.
Bottom Line: This resulted in tobacco having a haploid genome size of 4500 million base pairs, approximately four times larger than the related tomato (Solanum lycopersicum) and potato (Solanum tuberosum) genomes.We propose the combination of WGP physical mapping technology and tag profiling of ancestral lines as a generally applicable method to elucidate the ancestral origin of genome segments of polyploid species.The physical mapping of genes and their origins will enable application of biotechnology to polyploid plants aimed at accelerating and increasing the precision of breeding for abiotic and biotic stress resistance.
Affiliation: Biological System Research, Philip Morris International R&D, Philip Morris Products SA Quai Jeanrenaud 5, 2000, Neuchatel, Switzerland.Show MeSH
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Mentions: Sequence tags flanking EcoRI restriction sites for the whole genomes of N. sylvestris and N. tomentosiformis, the two closest relatives to the ancestral contributors of the S and T genomes to tobacco, were obtained by digesting genomic DNA of these lines using EcoRI and MseI, followed by amplification and Genome Analyzer II sequencing using the same method as applied for WGP map construction of N. tabacum. After quality filtering, 1 089 317 N. sylvestris and 1 035 343 N. tomentosiformis tags were obtained, compared with the 1 239 733 unique 51 nt tags obtained for N. tabacum during construction of the physical map, thus allowing determination of the ancestral origin of the tobacco tags, BACs and WGP physical map contigs by P value calculations for S or T tag enrichment. This analysis showed that 60.7% of the BACs are of S origin and 37.4% of T origin. The origin of 0.4% of the BACs could not be determined despite their S and/or T tags, and these BACs were classified as having an undefined origin. Finally, 1.5% of the BACs have no S or T tags and are thus of unknown origin. BACs consist of tobacco DNA fragments of approximately 100 kb, and as such should originate from only one of the two ancestors. Being able to assign an ancestral origin to 98.1% of the BACs indicates that S and T tags are not present randomly in BACs, thus validating the origin determination of WGP tags. The P value calculations for S or T tag enrichment further showed that 53.7% of the WGP contigs are of S origin, 45.8% are of T origin, 0.5% are of undefined origin, and only four are of unknown origin (Figure 1 and 2). Thus the S and T tags are rarely found together in the same WGP contig, providing additional confidence in the quality of the constructed physical map.
Affiliation: Biological System Research, Philip Morris International R&D, Philip Morris Products SA Quai Jeanrenaud 5, 2000, Neuchatel, Switzerland.