To date, one asparagine synthetase (AS) mRNA has been identified in Nicotiana tabacum. Our results point out that this gene encodes an actual AS protein. This proposal is supported by the following facts. (i) Blast analysis indicates that tobacco AS gene shows increased percent identity with several AS genes of different plant species. (ii) Leaf and root AS mRNA levels were inversely correlated with soluble carbohydrate concentrations, both in the presence of light and in darkness, and the addition of exogenous sucrose repressed the root AS gene expression. (iii) In light, a positive correlation between AS transcript levels and ammonium content was observed in both leaves and roots. (iv) A direct correlation between leaf AS gene expression and asparagine (Asn) concentration has been found. Furthermore, quantitative real-time PCR analyses showed that this gene expression was increased under boron (B) deficiency in roots, but not in leaves. Interestingly, soluble B concentration in roots of plants subjected to B deficiency was significantly lower than that of control plants. However, in leaves, there was no significant difference in soluble B content between both B treatments. We discuss the possible role of soluble B deficiency in the regulation of AS gene expression in tobacco roots. © 2010 Elsevier Ireland Ltd. All rights reserved.