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Type of phloem exudates and HPLC investigation out of amino acidic

Type of phloem exudates and HPLC investigation out of amino acidic

Plants from the field were harvested at the Cdos stage, screened for their tap root diameter (0.6–1.0 cm), before being acclimated in a climate-controlled room under hydroponic culture conditions as described in the previous section. Developmental stages were estimated from the phenological calendar established by the Bayer, BASF, Ciba and Hoechst companies. Nine plants for each developmental stage of the growth cycle were used for measuring HATS influx and BnNrt2 mRNA abundance. Six other plants were used for assaying amino acids content in phloem exudates.

Test step three (amino acid effects)

Six pots of 25 plants were first transferred to N-free nutrient solution for 12 h before being transferred to a new nutrient solution containing 1 m m KNO3 and either 100 µ m GABA, 1 m m GABA, 1 m m Gln, 1 m m Glu or 1 m m Asn for 12 h. At the end of the treatments, HATS influx, BnNrt2 mRNA abundance and root amino acids content were measured on six, two and three replicates, respectively.

Plant harvesting, nitrogen and you will isotope investigation

In order to measure nitrate influx, plants of the three experiments were grown in hydroponic tanks and were treated according to the particular experimental design. Roots were then washed twice in 1 m m CaSOcuatro solution for 1 minute at 20 °C to remove unabsorbed K 14 NO3 and transferred to influx solution containing 100 µ m of K 15 NO3 (99.9%) for 5 min. Plants were then washed twice in 1 m m CaSO4 solution for 1 min at 4 °C to slow down the metabolism and remove unabsorbed tracer residing in the cell wall space. Roots and shoots were harvested separately and dried for 48 h at 60 °C before being ground to fine powder for isotope analysis. (more…)

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