5 μL of purified fragments were transferred to a 384 printing plate together with 5 μL of 6 × SSC solution. The ESTs were printed onto poly-lysine coated slides with a Genemachines OmniGrid 100 (Genomic Solutions, Ann Arbor, MI, USA) using 16 SMP3 pins (TeleChem International, Sunnyvale, CA, USA). Printing conditions were 48% to 50% humidity and 21°C.
Support
glass
Coating
polysine
Description
Plant material for EST library construction: The Ein Avdat valley is located in the Negev desert in Israel at 450 m above sea level and a latitude of 30° 49' 30" N, and a longitude of 34° 46' 0" E. The average rainfall is 100 to 200 mm per year with most of the rain falling during winter time. Adult heart shaped leaves were collected from trees in areas A, B, and C for the cDNA library during the summer seasons of 2000 to 2001. Material was also harvested from young in vitro plantlets of a clone (clone B2) derived from a single tree from area B. Samples for microarray analysis were taken from areas A, B, and C and from the Ein Avdat parking lot (area P, 1 km from the valley) on 27 November 2003, and for ion, isotope and metabolite profiling on 27 November 2003 and 13 October 2004. The parking lot trees were irrigated with non saline tap water for 24 hours once a week. For normalized control libraries, leaves, shoots and roots were harvested from 8 month old P. euphratica seedlings. Seeds were obtained from the Forest Administration Division, Xinjiang Corps of Production and Construction, Urumqi, PR China. The seeds were sown on sand that had a high content of clay (6%) and germinated at 12 hours light with 250 μmol m-2 s-1 photosynthetically active radiation, 25°C, and a relative air humidity of 60% in a climate chamber (Weiss, Giessen, Germany). The seedlings were potted into soil (Fruhstorfer Erde N, Archut, Germany), transferred to a greenhouse with natural light and watered daily with Long Ashton macronutrient solution [52]. For stress treatments, leaves and roots were harvested from P. euphratica subjected to the following treatments: elevated CO2 (plants submitted to an atmosphere of 700 ppm for 60 days); different irradiance levels (plants under three different irradiance regimes corresponding to 48%, 18% and 8% external irradiance) for 60 days; drought stress (plants submitted to a 21 day cycle of drought with 5% to 3% volumetric soil humidity) before harvesting; flooding stress (plants submitted during 21 days to a complete submergence of the root system); ozone (plants submitted to 200 nl l-1 of O3 for 7 and 20 h); cold and freezing (leaves harvested from plants acclimated at 5 days of +2°C and from plants exposed to two and seven days of -2°C following the acclimation period); salt stress (leaves and xylogenic cambium harvested from plants exposed to 150 mM NaCl for 0.5 h, 1 h, 1.5 h, 2 h, 5 h, 10 h, 24 h and 48 h and roots harvested after 40 minutes, 1.5 h, 8 h and 20 h); cadmium stress (plants were stressed with 50 μM CdCl2 and roots harvested after 12, 24 and 48 h). Above text copied from Brosche et al. (2005) Genome Biology 6:R101 doi:10.1186/gb-2005-6-12-r101 PMID: 16356264
