Hani Al-Ahmad

Plants to power: bioenergy to fuel the future

Bioenergy should play an essential part in reaching targets to replace petroleum-based transportation fuels with a viable alternative, and in reducing long-term carbon dioxide emissions, if environmental and economic sustainability are considered carefully. Here, we review different platforms, crops, and biotechnology-based improvements for sustainable bioenergy. Among the different platforms, there are two obvious advantages to using lignocellulosic biomass for ethanol production: higher net energy gain and lower production costs. However, the use of lignocellulosic ethanol as a viable alternative to petroleum-based transportation fuels largely depends on plant biotechnology breakthroughs. We examine how biotechnology, such as lignin modification, abiotic stress resistance, nutrition usage, in planta expression of cell wall digestion enzymes, biomass production, feedstock establishment, biocontainment of transgenes, metabolic engineering, and basic research, can be used to address the challenges faced by bioenergy crop production.

Genetic load and transgenic mitigating genes in transgenic Brassica rapa (field mustard) × Brassica napus (oilseed rape) hybrid populations.

BackgroundOne theoretical explanation for the relatively poor performance of Brassica rapa (weed) × Brassica napus (crop) transgenic hybrids suggests that hybridization imparts a negative genetic load. Consequently, in hybrids genetic load could overshadow any benefits of fitness enhancing transgenes and become the limiting factor in transgenic hybrid persistence. Two types of genetic load were analyzed in this study: random/linkage-derived genetic load, and directly incorporated genetic load using a transgenic mitigation (TM) strategy. In order to measure the effects of random genetic load, hybrid productivity (seed yield and biomass) was correlated with crop- and weed-specific AFLP genomic markers. This portion of the study was designed to answer whether or not weed × transgenic crop hybrids possessing more crop genes were less competitive than hybrids containing fewer crop genes. The effects of directly incorporated genetic load (TM) were analyzed through transgene persistence data. TM strategies are proposed to decrease transgene persistence if gene flow and subsequent transgene introgression to a wild host were to occur.ResultsIn the absence of interspecific competition, transgenic weed × crop hybrids benefited from having more crop-specific alleles. There was a positive correlation between performance and number of B. napus crop-specific AFLP markers [seed yield vs. marker number (r = 0.54, P = 0.0003) and vegetative dry biomass vs. marker number (r = 0.44, P = 0.005)]. However under interspecific competition with wheat or more weed-like conditions (i.e. representing a situation where hybrid plants emerge as volunteer weeds in subsequent cropping systems), there was a positive correlation between the number of B. rapa weed-specific AFLP markers and seed yield (r = 0.70, P = 0.0001), although no such correlation was detected for vegetative biomass. When genetic load was directly incorporated into the hybrid genome, by inserting a fitness-mitigating dwarfing gene that that is beneficial for crops but deleterious for weeds (a transgene mitigation measure), there was a dramatic decrease in the number of transgenic hybrid progeny persisting in the population.ConclusionThe effects of genetic load of crop and in some situations, weed alleles might be beneficial under certain environmental conditions. However, when genetic load was directly incorporated into transgenic events, e.g., using a TM construct, the number of transgenic hybrids and persistence in weedy genomic backgrounds was significantly decreased.

Gene expression profiling of resistant and susceptible soybean lines infected with soybean cyst nematode

Soybean cyst nematode (SCN) is the most devastating pathogen of soybean. Information about the molecular basis of soybean–SCN interactions is needed to assist future development of effective management tools against this pathogen. Toward this end, soybean transcript abundance was measured using the Affymetrix Soybean Genome Array in a susceptible and a resistant reaction of soybean to SCN infection. Two genetically related soybean sister lines TN02-226 and TN02-275, which are resistant and susceptible, respectively, to the SCN race 2 infection were utilized in these experiments. Pairwise comparisons followed by false discovery rate analysis indicated that the expression levels of 162 transcripts changed significantly in the resistant line, of which 84 increased while 78 decreased. However, in the susceptible line, 1,694 transcripts changed significantly, of which 674 increased while 1,020 decreased. Comparative analyses of these transcripts indicated that a total of 51 transcripts were in common between resistance and susceptible responses. In this set, 42 transcripts increased in the resistant line, but decreased in the susceptible line. Quantitative real-time reverse-transcription polymerase chain reaction confirmed the results of microarray analysis. Of the transcripts to which a function could be assigned, genes were associated with metabolism, cell wall modification, signal transduction, transcription, and defense. Microarray analyses examining two genetically related soybean lines against the same SCN population provided additional insights into the specific changes in gene expression of a susceptible and a resistant reaction beneficial for identification of genes involved in defense.

Within-plant distribution and emission of sesquiterpenes from Copaifera officinalis

Copaifera officinalis, the diesel tree, is known for massive production of oleoresin, mainly composed of sesquiterpene hydrocarbons. In this study, composition of these sesquiterpenes and their concentrations in leaves, stems and roots of C. officinalis at two developmental stages, including the three-week old (TW) seedlings and two-year old (TY) trees, were determined. The leaves of TW seedlings and TY trees contained similar number of sesquiterpenes, which also had comparable concentrations. The stems of TW seedlings had higher concentrations of sesquiterpenes than those of TY trees. In contrast, the number of sesquiterpene species and their concentrations in the roots of TW seedlings were much lower than those in the roots of TY trees. Cluster analysis of sesquiterpenes estimated that there are at least four terpene synthase genes involved in the production of sesquiterpenes in C. officinalis. Because sesquiterpenes are highly volatile, emissions of sesquiterpenes from healthy and wounded TW seedlings were examined using headspace analysis. Whereas very low emission of sesquiterpenes was detected from undamaged plants, the physically injured seedlings emitted a large number of sesquiterpenes, the quality and the relative quantity of which were similar to those in leaves determined using organic extraction. The implications of our findings to the biosynthetic pathways leading to the production of sesquiterpenes as well as their biological roles in C. officinalis are discussed.

Assessing and Managing Biological Risks of Plants Used for Bioremediation, Including Risks of Transgene Flow

Abstract The plants used for phytoremediation pose special biological risks, whether transgenic or not, as most of the species: (a) are semi-domesticated; (b) are introduced from other habitats; (c) can become established in the contaminated site; (d) can spread and displace native species, and/or; (e) may introgress transgenes into related species. The addition of transgenes can reduce the risks, e.g. to sterilize or render the species and hybrid offspring hypersensitive to environmental effects (heat, cold), or to a chemical that will cull the species. Various measures can contain transgenes used in phytoremediation species to prevent gene flow, but most containment technologies are both uni-directional (prevent either outflow or influx), and are inherently leaky, even a concept specifically utilizable for phytoremediation Ð grafting non-transgenic scions on bioremediating transgenic rootstocks. Containment mechanisms should be either stacked with each other or with “mitigator” genes. Transgenic mitigation (TM) has mitigator genes added in tandem to the desired primary transgene (genetically linked) and the mitigator genes confer traits that are positive or neutral to the desired species but are deleterious to hybrids, keeping them at very low frequencies. The concept was demonstrated in tobacco and oilseed rape with a dwarfing mitigator gene that enhanced the reproductive productivity (harvest index) when cultured alone, but eliminated it from mixed populations. Besides the mitigator genes previously proposed for crop species (sterility, no seed shattering, dwarfing, no secondary dormancy) there are genes especially appropriate for phytoremediation, e.g. overexpression of cytokinin oxidase (reduces cytokinin levels) conferring reduced shoot systems (unfitness to compete) with a more extensive root system that is better for extracting toxic wastes as well as no-flowering for vegetatively propagated species. Thus, biotechnology can be harnessed to reduce risks from both non-transgenic and transgenic phytoremediation species.

Switchgrass (Panicum virgatum L.) cell suspension cultures: Establishment, characterization, and application

Switchgrass (Panicum virgatum L.) is a warm-season perennial grass that has received considerable attention as a potential dedicated biofuel and bioproduct feedstock. Genetic improvement of switchgrass is needed for better cellulosic ethanol production, especially to improve cellulose-to-lignin ratios. Cell suspension cultures offer an in vitro system for mutant selection, mass propagation, gene transfer, and cell biology. Toward this end, switchgrass cell suspension cultures were initiated from embryogenic callus obtained from genotype Alamo 2. They have been established and characterized with different cell type morphologies: sandy, fine milky, and ultrafine cultures. Characterization includes histological analysis using scanning electron microscopy, and utility using protoplast isolation. A high protoplast isolation rate of up to 10(6) protoplasts/1.0g of cells was achieved for the fine milky culture, whereas only a few protoplasts were isolated for the sandy and ultrafine cultures. These results indicate that switchgrass cell suspension type sizably impacts the efficiency of protoplast isolation, suggesting its significance in other applications. The establishment of different switchgrass suspension culture cell types provides the opportunity to gain insights into the versatility of the system that would further augment switchgrass biology research.

Biotechnology for bioenergy dedicated trees: meeting future energy demands

Abstract With the increase in human demands for energy, purpose-grown woody crops could be part of the global renewable energy solution, especially in geographical regions where plantation forestry is feasible and economically important. In addition, efficient utilization of woody feedstocks would engage in mitigating greenhouse gas emissions, decreasing the challenge of food and energy security, and resolving the conflict between land use for food or biofuel production. This review compiles existing knowledge on biotechnological and genomics-aided improvements of biomass performance of purpose-grown poplar, willow, eucalyptus and pine species, and their relative hybrids, for efficient and sustainable bioenergy applications. This includes advancements in tree in vitro regeneration, and stable expression or modification of selected genes encoding desirable traits, which enhanced growth and yield, wood properties, site adaptability, and biotic and abiotic stress tolerance. Genetic modifications used to alter lignin/cellulose/hemicelluloses ratio and lignin composition, towards effective lignocellulosic feedstock conversion into cellulosic ethanol, are also examined. Biotech-trees still need to pass challengeable regulatory authorities’ processes, including biosafety and risk assessment analyses prior to their commercialization release. Hence, strategies developed to contain transgenes, or to mitigate potential transgene flow risks, are discussed.