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Abstract Background The goals of our study were to assess the phylogeny and the population structure of tobacco accessions representing a wide range of genetic diversity; identify a subset of accessions as a core collection capturing most of the existing genetic diversity; and estimate, in the tobacco core collection, the extent of linkage disequilibrium (LD) in seven genomic regions using simple sequence repeat (SSR) markers. To this end, a collection of accessions were genotyped with SSR markers. Molecular diversity was evaluated and LD was analyzed across seven regions of the genome. Results A genotyping database for 312 tobacco accessions was profiled with 49 SSR markers. Principal Coordinate Analysis (PCoA) and Bayesian cluster analysis revealed structuring of the tobacco population with regard to commercial classes and six main clades were identified, which correspond to "Oriental", Flue-Cured", "Burley", "Dark", "Primitive", and "Other" classes. Pairwise kinship was calculated between accessions, and an overall low level of co-ancestry was observed. A set of 89 genotypes was identified that captured the whole genetic diversity detected at the 49 loci. LD was evaluated on these genotypes, using 422 SSR markers mapping on seven linkage groups. LD was estimated as squared correlation of allele frequencies (r2). The pattern of intrachromosomal LD revealed that in tobacco LD extended up to distances as great as 75 cM with r2 > 0.05 or up to 1 cM with r2 > 0.2. The pattern of LD was clearly dependent on the population structure. Conclusions A global population of tobacco is highly structured. Clustering highlights the accessions with the same market class. LD in tobacco extends up to 75 cM and is strongly dependent on the population structure.

Leaves of tobacco ( Nicotiana tabacum) are covered with glandular trichomes that produce sucrose esters and diterpenoids in varying quantities, depending on cultivar type. The bicyclic diterpene Z-abienol is the major labdanoid present in some oriental tobacco cultivars, where it constitutes a precursor of important flavours and aromas. We describe here the identification and characterization of two genes governing the biosynthesis of Z-abienol in N. tabacum. As for other angiosperm labdanoid diterpenes, the biosynthesis of Z-abienol proceeds in two steps. NtCPS2 encodes a class-II terpene synthase that synthesizes 8-hydroxy-copalyl diphosphate, and NtABS encodes a kaurene synthase-like (KSL) protein that uses 8-hydroxy-copalyl diphosphate to produce Z-abienol. Phylogenetic analysis indicates that NtABS belongs to a distinct clade of KSL proteins that comprises the recently identified tomato ( Solanum habrochaites) santalene and bergamotene synthase. RT-PCR results show that both genes are preferentially expressed in trichomes. Moreover, microscopy of NtCPS2 promoter-GUS fusion transgenics demonstrated a high specificity of expression to trichome glandular cells. Ectopic expression of both genes, but not of either one alone, driven by a trichome-specific promoter in transgenic Nicotiana sylvestris conferred Z-abienol formation to this species, which does not normally produce it. Furthermore, sequence analysis of over 100 tobacco cultivars revealed polymorphisms in NtCPS2 that lead to a prematurely truncated protein in cultivars lacking Z-abienol, thus establishing NtCPS2 as a major gene controlling Z-abienol biosynthesis in tobacco. These results offer new perspectives for tobacco breeding and the metabolic engineering of labdanoid diterpenes, as well as for structure-function relationship studies of terpene synthases. [ABSTRACT FROM AUTHOR]

Using single-copy conserved ortholog set (COSII) and simple sequence repeat (SSR) markers, we have constructed two genetic maps for diploid Nicotiana species, N. tomentosiformis and N. acuminata, respectively. N. acuminata is phylogenetically closer to N. sylvestris than to N. tomentosiformis, the latter two of which are thought to contribute the S-genome and T-genome, respectively, to the allotetraploid tobacco ( N. tabacum L., 2 n = 48). A comparison of the two maps revealed a minimum of seven inversions and one translocation subsequent to the divergence of these two diploid species. Further, comparing the diploid maps with a dense tobacco map revealed that the tobacco genome experienced chromosomal rearrangements more frequently than its diploid relatives, supporting the notion of accelerated genome evolution in allotetraploids. Mapped COSII markers permitted the investigation of Nicotiana–tomato syntenic relationships. A minimum of 3 (and up to 10) inversions and 11 reciprocal translocations differentiate the tomato genome from that of the last common ancestor of N. tomentosiformis and N. acuminata. Nevertheless, the marker/gene order is well preserved in 25 conserved syntenic segments. Molecular dating based on COSII sequences suggested that tobacco was formed 1.0MYA or later. In conclusion, these COSII and SSR markers link the cultivated tobacco map to those of wild diploid Nicotiana species and tomato, thus providing a platform for cross-reference of genetic and genomic information among them as well as other solanaceous species including potato, eggplant, pepper and the closely allied coffee (Rubiaceae). Therefore they will facilitate genetic research in the genus Nicotiana. [ABSTRACT FROM AUTHOR]

Senescence is a genetically controlled mechanism that modifies leaf chemistry. This involves significant changes in the accumulation of carbon- and nitrogen-containing compounds, including asparagine through the activity of asparagine synthetases. These enzymes are required for nitrogen re-assimilation and remobilization in plants; however, their mechanisms are not fully understood. Here, we report how leaf curing—a senescence-induced process that allows tobacco leaves to dry out—modifies the asparagine metabolism. We show that leaf curing strongly alters the concentration of the four main amino acids, asparagine, glutamine, aspartate, and glutamate. We demonstrate that detached tobacco leaf or stalk curing has a different impact on the expression of asparagine synthetase genes and accumulation of asparagine. Additionally, we characterize the main asparagine synthetases involved in the production of asparagine during curing. The expression of ASN1 and ASN5 genes is upregulated during curing. The ASN1-RNAi and ASN5-RNAi tobacco plant lines display significant alterations in the accumulation of asparagine, glutamine, and aspartate relative to wild-type plants. These results support the idea that ASN1 and ASN5 are key regulators of asparagine metabolism during leaf curing. [ABSTRACT FROM AUTHOR]