It is still unclear what parameter(s), other than grain yield, might be a suitable indicator in a wheat (Triticum aestivum L.) breeding program for drought resistance. In this study, the leaf relative water content (RWC) and gas‐exchange parameters were compared between a drought‐resistant winter wheat genotype (cv. TAM W‐101) and a drought‐susceptible genotype (cv. Sturdy) to determine if these physiological parameters contribute to drought resistance in TAM W‐101. Plants were grown under well‐watered conditions in growth chambers until drought stress was imposed by limited watering of plants at anthesis or during vegetative growth. In both growth stages, TAM W‐101 maintained a higher RWC and apparent photosynthesis (A) than Sturdy under moderate to severe drought stress. TAM W‐101 plants also maintained a higher photosynthetic capacity (higher A at a given intercellular CO2 concentration [Ci]) under stress than did Sturdy in both growth stages. Photosynthetic water use efficiency (pWUE = A/stomatal conductance) generally increased with stress severity until very severe stress levels were attained. Thus, genotypic pWUE comparisons using stressed plants should be evaluated on a water‐status basis (e.g., RWC) to avoid the confounding effect of stress severity on pWUE. TAM W‐101 tended to have higher pWUE (RWC basis) than Sturdy under moderate to severe stress conditions, but not under well‐watered conditions. High leaf RWC, A, and photosynthetic capacity are traits that may contribute to drought resistance in TAM W‐101.

To more fully understand the biochemical and molecular events which occur in plants exposed to nitrate, cDNAs whose accumulation was enhanced in nitrate- and cycloheximide-treated maize (Zea mays L. W64A x W182E) roots were isolated. The 340 bp Zmrprn1 (for Zea mays root primary response to nitrate) cDNA also hybridized with a probe enriched for nitrate-induced sequences, and was characterized further. Sequence analysis of a near full-length cDNA (Zmrprn1A) showed strong homology (> 90% amino acid identity) with a root ferredoxin-NADP+ oxidoreductase (FNR) of rice, and 45-50% amino acid identify with leaf FNR genes. When expressed in Escherichia coli, the Zmrprn1A cDNA produced a protein with NADPH: ferricyanide reductase activity, consistent with the enzymatic properties of an FNR. The Zmrprn1 cDNA hybridized with a 1.4 kb transcript which was expressed in the maize root primary response to nitrate. That is, mRNA levels in roots increased rapidly and transiently in response to external nitrate, and low levels of nitrate (10 microM) induced transcript accumulation. The accumulation of the Zmrprn1 transcript was not prevented by cycloheximide, indicating that the cellular factor(s) required for expression were constitutively present in maize roots. The Zmrprn1 mRNA accumulated specifically in response to nitrate, since neither K+ nor NH4+ treatment of roots caused transcript accumulation. Maize leaves had about 5% of the transcript level found in roots, indicating a strong preference for expression of Zmrprn1 in roots. Analysis of maize genomic DNA indicated the presence of only a single gene or very small gene family for the Zmrprn1. Together, the data indicate that Zmrprn1A encodes a nitrate regulated maize root FNR.

Transient expression of GUS in rice (Oryza sativa L.) mediated by Agrobacterium tumefaciens was characterized using binary vectors containing gusA genes that express minimal (pKIWI105 and pCNL1) or no (p35S-GUS-INT and pCNL56) GUS activity in bacteria. Four-day old seedlings obtained from seeds or immature embryos of rice were cut into shoot, root, and seed remnants and inoculated with various strains of A. tumefaciens. Transient GUS expression events were quantitated histochemically by determining the frequency of explants exhibiting blue spots indicative of GUS at four to six days after cocultivation with A. tumefaciens. A. tumefaciens strains that did not contain the gusA gene (At643) or a Ti-plasmid (At563 and At657) did not elicit any blue staining characteristic of GUS activity. Several parameters were important in obtaining efficient transient expression of GUS in rice mediated by A. tumefaciens. The growth regulator 2,4-D inhibited GUS expression if present during the seed germination period, but the presence of 6 mg/l 2,4-D during cocultivation of the explants with A. tumefaciens slightly enhanced GUS expression efficiency. All 21 rice cultivars tested expressed GUS after co-cultivation with A. tumefaciens. The GUS expression frequency was highest amongst the indica cultivars. The frequencies of GUS expression in japonica cultivars and in Oryza glaberrima cultivars (grown primarily in Africa) were generally one-half to one-third the level found for indica varieties. Leaf explants were more susceptible to A. tumefaciens-facilitated GUS expression than were roots or seed remnants. The vir genes of an agropine-type Ti-plasmid of A. tumefaciens were most effective in directing transient GUS expression in rice, whereas those of a nopaline-type and an octopine-type plasmid were less effective. We have also found that the frequency of transient expression of GUS was higher with pBIN19 as the precursor cloning vector than with pEND4K as the precursor cloning vector. Reasons for differences in effectiveness of these binary vectors are discussed. Using the conditions described here, A. tumefaciens-mediated frequencies of transient GUS expression in four-day old shoots of several rice cultivars were routinely in excess of 50%.

An efficient co-transformation protocol using polyethylene glycol was developed for Zea mays L. (cv. A188 x BMS) protoplasts isolated from suspension culture cells. Co-transformation was accomplished by using plasmid constructions containing beta-glucuronidase (gusA) or neomycin phosphotransferase (neo) gene coding sequences; both were under control of the CaMV 35S promoter. Protoplast culture and transformation conditions were optimized to assure efficient recovery of transformed cells. The overall efficiency of transformation was 1 x 10(-4) (calculated per viable protoplast plated). Among kanamycin-resistant lines, 50% showed a high level of GUS activity (above one unit). Southern blot hybridization confirmed the presence of numerous gusA and neo coding sequences in the maize genome. In two analyzed lines, integrated sequences appeared to be organized in tandem head-to-tail repeats. Results also indicated that the integrated sequences were partially methylated.