Effects of water stress and constitutive expression of a drought induced chitinase gene on water-use efficiency and carbon isotope composition of strawberry

Recombinant DNA technology and tracer technique are being widely used for different purposes. Plant genetic transformation is a common practice for increasing crop tolerance to biotic and abiotic stresses and stable isotopes is a good tool to track the effects of environmental stresses on crop productivity. In this research, the effect of water stress on plant carbon isotopic composition (δ13C) and water use efficiency (WUE), as indicators of physiological processes and environmental factors, were evaluated for transgenic strawberry (Fragaria x ananassa) plants continuously expressing a chitinase gene and non-transgenic plants. To implement water stress, strawberry plants were grown in pots and were assigned to three different soil water contents (SWC) of well-watered, moderately-watered and water-stressed. Treatments were implemented for two months, in a growth chamber. At the final stage of the experiment, leaves were randomly collected from each experimental unit. After oven-drying and powdering, subsamples of 4 to 5 mg of plant materials were combusted under vacuum, using Vycor tubes containing cupric oxide and silver wire. Combustion took place at 520 degrees C for 5 hrs to produce carbon dioxide. Carbon isotope composition (δ13C) was measured relative to Vienna Pee Dee Belmnite (VPDB). Both SWC and the expression of chitinase gene in transgenic strawberry plants significantly affected δ13C and WUE (p < 0.001). Highly significant (p < 0.01), but negative correlations were found between δ13C and plant dry matter components, such as lead dry matter (r = -0.92; p = 0.009) and total dry matter (r = -0.92; p = 0.008). High correlations were found between δ13C and water use per plant (r = -0.93; p = 0.006), and between δ13C and leaf area per plant (r = -0.93; p = 0.006)

Expression of the β-1,3-glucanase gene bgn13.1 from Trichoderma harzianum in strawberry increases tolerance to crown rot diseases but interferes with plant growth

The expression of antifungal genes from Trichoderma harzianum, mainly chitinases, has been used to confer plant resistance to fungal diseases. However, the biotechnological potential of glucanase genes from Trichoderma has been scarcely assessed. In this research, transgenic strawberry plants expressing the β-1,3-glucanase gene bgn13.1 from T. harzianum, under the control of the CaMV35S promoter, have been generated. After acclimatization, five out of 12 independent lines analysed showed a stunted phenotype when growing in the greenhouse. Moreover, most of the lines displayed a reduced yield due to both a reduction in the number of fruit per plant and a lower fruit size. Several transgenic lines showing higher glucanase activity in leaves than control plants were selected for pathogenicity tests. When inoculated with Colletotrichum acutatum, one of the most important strawberry pathogens, transgenic lines showed lower anthracnose symptoms in leaf and crown than control. In the three lines selected, the percentage of plants showing anthracnose symptoms in crown decreased from 61 % to a mean value of 16.5 %, in control and transgenic lines, respectively. Some transgenic lines also showed an enhanced resistance to Rosellinia necatrix, a soil-borne pathogen causing root and crown rot in strawberry. These results indicate that bgn13.1 from T. harzianum can be used to increase strawberry tolerance to crown rot diseases, although its constitutive expression affects plant growth and fruit yield. Alternative strategies such as the use of tissue specific promoters might avoid the negative effects of bgn13.1 expression in plant performance.This research was supported by the Comisión Interministerial de Ciencia y Tecnología in Spain and FEDER EU Funds (Grant No. AGL2011-30354-C02-01).Peer reviewe