Plant microRNAs have been implicated in various abiotic stress responses. We identified several conserved microRNAs that showed differential expression in Medicago truncatula plants subjected to water deficit: miR169 is down-regulated only in the roots and miR398a/b and miR408 are strongly up-regulated in both shoots and roots. Down-regulation of miR169 in the roots did not correlate with accumulation of its target MtHAP2-1 transcripts, suggesting that its regulation may not occur at the mRNA level or may depend on other regulatory mechanisms, which do not involve this miRNA, in water-deficit conditions. The up-regulation of miR398a/b and miR408 and the clear down-regulation of their respective target genes, which encode the copper proteins COX5b (subunit 5b of mitochondrial cytochrome c oxidase) and plantacyanin, highlight the involvement of these miRNAs in response to water deprivation in M. truncatula. Also, miR398 up-regulation is inversely correlated with the down-regulation of copper superoxide dismutase, CSD1, during water deficit. The regulation of genes encoding copper proteins by miR398a/b and miR408 suggests a link between copper homeostasis and M. truncatula adaptation to progressive water deficit.

Background: High-throughput sequencing technology is capable to identify novel short RNAs in plant species. We used Solexa sequencing to find new microRNAs in one of the model legume species, barrel medic (Medicago truncatula).

Trehalose (a non-reducing disaccharide) plays an important role in abiotic stress protection. It has been shown that using trehalose synthesis genes of bacterial origin, drought and salt tolerance could be achieved in several plants. A cassette harboring the AtTPS1 gene under the control of the CaMV35S promoter and the Bialaphos resistance gene was inserted in the binary plasmid vector pGreen0229 and used for Agrobacterium-mediated transformation of tobacco (Nicotiana tabacum). T0 plants obtained were analyzed by PCR for the presence of AtTPS1 gene. Thirty lines were positive and seeds were germinated on media with 6 mg/l PPT to obtain T1 plants that were grown in the greenhouse to obtain T2 seeds that were germinated on selective media. Lines which seeds showed a 100 % survival rate were considered homozygous transgenic T1 lines. Three lines were selected and gene expression confirmed by northern and western blots. Transgenic seeds were germinated on media with different concentrations of mannitol (0, 0.25, 0.5 and 0.75 M) and sodium chloride (0, 0.07, 0.14, 0.2, 0.27 and 0.34 M) to score their tolerance to osmotic stress. Assays were conducted to test the tolerance of transgenic plants to drought (measurement of water percentage as a consequence of water withdrawal), desiccation (measurement of water loss as a consequence leaf detaching) and temperature stresses (germination at 15 • C and 35 • C). Transgenic tobacco plant lines registered higher germination rates under osmotic and temperature stress situations than did wild-type plants. Responses to drought and desiccation stresses were similar for all plant lines. It can hence be suggested that the heterologous expression of TPS1 gene from Arabidopsis can be used successfully to increase abiotic stress tolerance in model plants and probably in other crops.

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