Research projects


Program Responsible: Georgina Hernández-Delgado, Ph.D.

Research Projects

  • Functional genomics of common bean (Phaseolus vulgaris).
  • Other ongoing related project of this program include a) the molecular and physiological characterization of transgenic model legumes altered in nodule nitrogen assimilation and b) the exploration of DNA transfer to plants by Rhizobium etli .

Subprojects


Top Functional genomics of common bean (Phaseolus vulgaris).

Common beans ( Phaseolus vulgaris L.) are the world's most important grain legume for direct human consumption. It is widely produced specially in Latin America and Africa . In Mexico , and other countries of Central and South America , beans are staple crops serving as the primary source of protein nitrogen (N) in the diet. In sustainable agricultural practice, beans and other legumes play a significant role in effective management of fertilizer, improving soil health, and protection of surface and ground water from contamination.

Production of high-quality, nutritious food requires N and phosphorus (P). However, most of the Latin American soils are poor in N and/or are acid. Acidic soils are characterized by their low capacity for cationic exchange and, therefore, commonly exhibit deficiency in mineral nutrients such as P, calcium, magnesium, zinc, as well as enhance aluminum (Al) and manganese toxicity. Use of beans and other legumes on farms can help remedy both overuse of N and P fertilizers in wealthy countries and insufficient sources of the inputs in poorer countries. Legumes can produce their own N fertilizer through a process known as symbiotic N 2 fixation. This process occurs through the symbiotic association between legumes and soil bacteria known as rhizobia. In addition, legumes can be fairly efficient at acquiring soil P, thereby requiring less P fertilizer.

The advent of genomic research in plants has increased, dramatically, our knowledge of global plant biology. Genomic and post-genomic approaches -so called ¡ omics: transcriptomics, proteomics, metabolomics- in legume research have lead to the discovery of hundreds of new genes that appear to participate in different processes such as plant-microbe interactions and nutrient acquisition. Large scale EST (expressed sequence tags) sequencing has provided us with in-depth overview of the nodule trabscriptome especially in the model legume Medicago truncatula .

Despite the importance of common beans as crop legumes, few genomic resources exist for this species. Scientists from this program have performed collaborative research with Dr Carroll P. Vance, from U. Minnesota/USDA, with the aim of developing molecular tools and generating resources to build the foundations for functional genomics of beans. The resources generated in this collaborative project include: a) Four cDNA libraries, from mRNA isolated from Phaseolus: root nodules, P-deficiency stressed roots, developing and mature pods, and leaves, b) Sequence of some 20,000 ESTs from those bean organs. Our contribution, together with ca. 5000 ESTs sequences provided by Melotto et al., have generated a common bean Gene Index of 9,484 unique gene set, comprised by 2,906 TC sequences and 6,578 singleton ESTs (The Institute of Genomic Research). In addition, high-density macroarrays (nylon membranes) with some 3000 ESTs from each of the cDNA libraries have been printed, to perform transcriptome expression studies. Bean root nodule transcript profile revealed several genes that are most highly expressed in bean nodules as compared to root, leaf and pod; these include those gene that participate in key C/N metabolic pathways that lead to ureide synthesis, which are the major nitrogen transporters in common bean.

Within this research line and based on the genomic resources already generated, current research in this program is centered in the functional genomics of nodulation/symbiotic nitrogen fixation and of adaptation to abiotic stresses: P starvation and metal toxicity. Following is a brief summary of the subproject that scientists, technicians and students of this program are currently investigating:


Top Regeneration and genetic transformation of Phaseolus vulgaris

Legumes are notoriously difficult to regenerate and transform, and, certainly, this is the case for bean. Though scientists from this program, as well as scientist from different countries, have tried extensively, there is still no reproducible and efficient protocol for bean genetic transformation. Current exploration in this program include: the use of different Phaseolus vulgaris genotypes to identify those with better regeneration capacity, which in turn can be used for Agrobacterium tumefaciens mediated transformation; the development of an efficient protocol for bean A. rhizogenes mediated transformation, similar as those reported for the model legumes Medicago truncatula and Lotus japonicus . These protocols rapidly and efficiently generate composite plants that contain a transgenic root system and a non-transgenic shoot.

Disruption of normal gene expression is a requisite to determining gene function. Effective gene silencing in plants has been achieved through the delivery of RNAi. The mechanism through which RNAi exerts its effect involves the creation and recognition of double-stranded RNA that triggers sequence-specific degradation of cellular transcripts. In this context, bean genetic transformation is crucial for the advancement of different project oriented to define specific gene function (see below).


Top Transcriptomics of nodule development

Scientist from this program in collaboration with scientists from U. Minnesota have reported the transcript profile of mature bean nodules elicited by R. tropici . Current projects, using macroarray membranes spotted with ca. 3000 ESTs from the nodule cDNA library, are oriented to define the transcript profile of nodules at different developmental stages (early, mature, senescent). Results from this project would allow to find new genes, to define key genes and metabolic/developmental pathways and to define regulatory networks of nodule ontogeny.


Top Functional genomics of bean-Rhizobium symbiotic interactions and nodule metabolism

The development of symbiotic nodules is initiated by complex interactions between rhizobial bacteria and legume plants. Nodulation is regulated by a two-way exchange of plant and bacterial signal molecules. Though genomic approaches have provided important information about key genes involved in these processes, especially in the model legumes, plant molecular mechanisms of rhizobial signal molecules (Nod factors) perception and of subsequence signal transduction and nodule organogenesis are still largely unknown. The nodulation process involves a drastic alteration of the expression of host legume genes. Comprehensive analysis of gene expression profiles during the nodulation process has critical importance in understanding Rhizobium -legume interactions, and subsequent nodule formation and metabolism. Results from this program derived from transcriptome macroarray analysis provided a comprehensive data source for investigation of molecular mechanisms underlying nodulation and symbiotic nitrogen fixation. Current research of this program includes the expressional analysis of novel nodule-enhanced signaling elements, and carbon and nitrogen metabolism genes together with promoter analysis of representative carbon/nitrogen metabolism genes. With the aim of defining specific roles and relevance of selected signal elements or metabolic genes, gene disruption approaches -RNAi- will be followed and is based in obtaining positive results from the project describe above (1). An alternative to unsuccessful bean transformation would be to use model legume systems such as Lotus japonicus with efficient genetic transformation protocols that have been used in this program.


Top Functional genomics of bean adaptation to abiotic stresses: P starvation and metal toxicity.

This program is initiating this project using the generated resource of spotted macroarray membranes with ca. 3000 ESTs from bean P starved roots (-P roots) cDNA library. The general goal is to assess global gene expression in bean roots, nodules and leaves in response to the abiotic stresses: P starvation and metal (Al and Mn) toxicity. Comparison of gene expression between various organs at different developmental stages and different growth conditions (stress) offer the possibility to determine temporal and spatial relationships in the hierarchy of functional and regulatory gene networks. This project will take advantage of the wide genetic variety of bean germplasm, by performing comparative studies among control varieties and varieties that have been characterized or selected as tolerant to the mentioned abiotic stresses. The identification of genes that contribute to plant adaptation to nutrient stress is an important element in developing germplasm that can grow deficiently in nutrient poor soils.

Other ongoing related project of this program include a) the molecular and physiological characterization of transgenic model legumes altered in nodule nitrogen assimilation and b) the exploration of DNA transfer to plants by Rhizobium etli.