USDA-ARS-Fargo Barley Genetics Laboratory members, summer 2007: Top Row - Hanna Fischer and Paige Scherer (NDSU Governor's School - High School students), Jim Hegvik (NDSU undergrad). Bottom Row - Neerja Tyagi (NDSU Plant Sciences PhD student), Lynn Dahleen, Bill Morgan (lab technician), Nittin Mittal (NDSU undergrad).

Muthusamy Manoharan at Plant biotechnology laboratory, University of Arkansas - Pine Bluff

Barley was first transformed using particle bombardment in the early 1990's by several labs and a very thorough review was published by Peggy Lemaux et al. in 1999, soon after success was achieved with Agrobacterium-mediated transformation. Since then, research has expanded across the world. My coauthor, Dr. Muthusamy Manoharan, joined my lab in 1999 as a North Dakota State University Postdoctoral Research Associate as we began our own barley transformation experiments. Together, we transformed barley with several antitoxin and antifungal genes to try to increase resistance to Fusarium head blight. When Manoharan moved on to his current position as Assistant Professor at the Department of Agriculture, University of Arkansas-Pine Bluff in 2002, we continued our collaboration, evaluating lines he developed before he left Fargo and getting his lab set up to do additional barley transformations. When Dr. Prakash Lakshmanan, editor for In Vitro-Plant, asked us if we would write a review of barley transformation research, we used the opportunity to describe the extensive research reported since Peggy's review in 1999. In this review, we cover advances in transformation methods, the challenges encountered in barley transformation, development of transformation systems for commercial cultivars, gene expression, stability and inheritance, and gene flow. The review also describes recent research on gene tagging through transformation, insertion of disease resistance and abiotic stress resistance genes, transformation with genes for improved malting quality, nutrient content, feed quality and production of feed enzymes and pharmaceutical compounds. I am constantly amazed by the breadth of research being conducted around the world and look forward to all the new reports to come. Lynn S. Dahleen and Muthusamy Manoharan. Recent Advances in Barley Transformation, In Vitro Cellular & Developmental Biology-Plant, 43: 493-506, 2007.

Left to right: Eldridge Wynn, TJ Evens, Peter D'Aiuto, Scott Hyndman, and Randy Niedz.

Regulating Plant Tissue Growth by Mineral Nutrition

Providing the correct types and concentrations of mineral nutrients is essential for achieving "good" in vitro growth. However, given that there are thirteen putatively essential mineral nutrients that probably exhibit significant interactive effects, determining the appropriate levels of these nutrients for any in vitro system is a nontrivial and complex experimental problem. To examine a possible approach to address this experimental complexity in a manner useful for plant tissue culture, we structured the problem into a geometric framework for experimental design efficiency and utility. This allowed us to sample a five-factor design space with 33 treatment combinations rather than the 243 required for a 3-level factorial. Though this approach identified a nutrient formulation that yielded a 40% increase in citrus callus growth, the most important result was the systematic efficiency of the methodology used to identify and characterize the resultant "high growth" formulation. The identified "high growth" formulation is not simply an isolated recipe, but rather a quantified region in the 5-factor experimental design space. We think it is now possible to envision the efficient characterization of any in vitro system for a seemingly complex array of abiotic/biotic parameters that previously have been quantified in a relatively disjointed manner. The result will be significantly improved growth responses derived from a deeper understanding of the role and interactions of all the components and conditions comprising complex in vitro systems. Randall P. Niedz and Terrence J. Evans. Regulating Plant Tissue Growth by Mineral Nutrition, In Vitro Cellular & Developmental Biology-Plant, 43: 370-382, 2007.

This manuscript will be discussed in more depth at the 2008 World Congress on In Vitro Biology. More information on the session is listed here under Plant Symposia Monday from 5:00 pm - 6:00 pm.

Left to right: Praveen K. Saxena and Chunzhao Liu

Echinacea Biotechnology: Challenges and Opportunities

Echinacea, better known as purple coneflower, is one of the most commonly used herbal remedy in North America and Europe. It has received global attention because of its potential application in a range of health conditions. Further, there is enormous potential for the discovery of new medicinal compounds in this species and an immediate need for techniques to facilitate the production of high quality, chemically consistent plant material for drug development and clinical trials. In vitro tissue culture of Echinacea can play a vital role in the development of novel germplasm, rapid multiplication, and genetic modifications for an enhanced phytochemical production. Recent establishment of liquid culture techniques, large-scale bioreactors and Agrobacterium-mediated transformation in our group is changing the production parameters of the Echinacea species. This review provides an overview of the recent developments in in vitro technologies and challenges that remain in the Echinacea biotechnology. Bilal Haider Abbasi, Praveen K. Saxena, Susan J. Murch, and Chun-Zhao Liu. Echinacea Biotechnology: Challenges and Opportunities, In Vitro Cellular and Developmental Biology - Plant, 43: 481-492, 2007.

Biotechnological Approaches for Parasitic Weed Control

Parasitic weeds (over 4000 species) represent one of the most destructive and intractable problems to agricultural production, causing considerable losses of 30-80% in staple food and industrial crops in the world. Parasitic weeds adopt different forms to invade host plants. Some (dodders and mistletoes) invade aerial parts, while others invade the underground roots (Orobanche and Striga). Parasitic plants vary widely in their degree of host-dependence. Some parasites are photosynthetic and have the ability to survive without a host, but are able to take advantage of an available host to augment their nutrition (facultative parasites i.e. Triphysaria spp.). Other parasites have an absolute host-requirement, but retain some photosynthetic capacity (obligate hemiparasites, i.e. Striga and Alectra spp., mistletoes and some Cuscuta spp.) The nature of parasitic weeds makes control extremely difficult, costly or hazardous to the environment. Although, the simplest and most effective approach to parasitic weeds control - host resistance - remains an unrealized goal for agriculture, a wide variety of parasitic weed control (chemical, biological, cultural and resistant crops) has been tried. Unfortunately, most are partially effective and have significant limitations. Development of new effective strategy for resistance to parasitic weeds requires 1) identification of genes whose products selectively inhibit parasite growth or 2) a target key-gene of the parasitic for silencing. Our group have generated transgenic tobacco plants expressing a cecropin peptide (sarcotoxin IA), under the inducible control of the HMG2 promoter. These plants while having no obvious effect on the host plant growth and development, showed enhanced resistance to the parasitic weed Orobanche. Following parasite inoculation transgenic lines, showed higher numbers of aborted parasitization events, reduced Orobanche biomass and increased host biomass, compared to non-transgenic controls. While achieving the first approach, the discovery of RNA silencing was one of the most important biotechnological findings of the last decade. Small interfering RNAs (siRNAs) are short double-stranded RNA molecules that facilitate potent and sequence-specific gene suppression by degradation of mRNA sequences to which they are homologous, thereby silencing the target gene. Parasitic weeds such as Orobanche accumulate high amounts of mannitol during their development. Mannitol content in the parasite is regulated by Mannose 6-Phosphate Reductase (M6PR), an essential process to Orobanche for water and nutrient uptake from the host. In our study, we used the inverted repeat technique for gene silencing of M6PR, a key-gene in Orobanche spp. in order to provide the host plant with resistance against the parasite. Our results showed that the endogenous M6PR mRNA from O. aegyptiaca tubercles or shoots grown on transgenic tomato plants harboring the M6PR silencing construct was reduced by 60-80%. The number of dead tubercles was also increased significantly on transgenic plants as compared with the control plants. In summary: Optimal parasitic weed control could be achieved by either the use of parasite-resistance crops (from conventional breeding), or by crops genetically engineered for resistance. o far only a few crop varieties with stable resistance have been developed after decades of conventional plant breeding, and genetic resources for resistance genes are limited. Therefore, alternative biotechnological approaches could be ideal for parasitic weed control. Advantages of these approaches are: reducing labor, lowering expense, increasing cropping choices, and elimination of the need for chemicals that may be harmful to the environment. Beside advantages, disadvantages could be aroused, since expression of novel genes in crop plants can pose food safety issue and there is a continuous concern to gene transfer from transgenic crop plants to wild plants. Radi Aly. Conventional and Biotechnological Approaches for Control of Parasitic Weeds, In Vitro Cellular and Developmental Biology - Plant, 43: 304-317, 2007.