Articles by Miguel Lara in JoVE
Plant Promoter Analysis: Identification and Characterization of Root Nodule Specific Promoter in the Common Bean Kalpana Nanjareddy*1, Manoj-Kumar Arthikala*1, Alma-Leticia Aguirre1, Brenda-Mariana Gómez1, Miguel Lara2 1Ciencias Agrogenómicas, Escuela Nacional de Estudios Superiores Unidad León- Universidad Nacional Autónoma de México (UNAM), 2Instituto de Biología, Universidad Nacional Autónoma de México (UNAM), Ciudad Universitaria, Coyoacan Promoter expression analyses are crucial to improving the understanding of gene regulation and the spatiotemporal expression of target genes. Herein we present a protocol to identify, isolate, and clone a plant promoter. Further, we describe the characterization of the nodule-specific promoter in the common bean hairy roots.
Other articles by Miguel Lara on PubMed
Protoplast Isolation, Transient Transformation of Leaf Mesophyll Protoplasts and Improved Agrobacterium-mediated Leaf Disc Infiltration of Phaseolus Vulgaris: Tools for Rapid Gene Expression Analysis BMC Biotechnology. | Pubmed ID: 27342637 Phaseolus vulgaris is one of the most extensively studied model legumes in the world. The P. vulgaris genome sequence is available; therefore, the need for an efficient and rapid transformation system is more imperative than ever. The functional characterization of P. vulgaris genes is impeded chiefly due to the non-amenable nature of Phaseolus sp. to stable genetic transformation. Transient transformation systems are convenient and versatile alternatives for rapid gene functional characterization studies. Hence, the present work focuses on standardizing methodologies for protoplast isolation from multiple tissues and transient transformation protocols for rapid gene expression analysis in the recalcitrant grain legume P. vulgaris.
A Legume TOR Protein Kinase Regulates Rhizobium Symbiosis and Is Essential for Infection and Nodule Development Plant Physiology. | Pubmed ID: 27698253 The target of rapamycin (TOR) protein kinase regulates metabolism, growth, and life span in yeast, animals, and plants in coordination with nutrient status and environmental conditions. The nutrient-dependent nature of TOR functionality makes this kinase a putative regulator of symbiotic associations involving nutrient acquisition. However, TOR's role in these processes remains to be understood. Here, we uncovered the role of TOR during the bean (Phaseolus vulgaris)-Rhizobium tropici (Rhizobium) symbiotic interaction. TOR was expressed in all tested bean tissues, with higher transcript levels in the root meristems and senesced nodules. We showed TOR promoter expression along the progressing infection thread and in the infected cells of mature nodules. Posttranscriptional gene silencing of TOR using RNA interference (RNAi) showed that this gene is involved in lateral root elongation and root cell organization and also alters the density, size, and number of root hairs. The suppression of TOR transcripts also affected infection thread progression and associated cortical cell divisions, resulting in a drastic reduction of nodule numbers. TOR-RNAi resulted in reduced reactive oxygen species accumulation and altered CyclinD1 and CyclinD3 expression, which are crucial factors for infection thread progression and nodule organogenesis. Enhanced expression of TOR-regulated ATG genes in TOR-RNAi roots suggested that TOR plays a role in the recognition of Rhizobium as a symbiont. Together, these data suggest that TOR plays a vital role in the establishment of root nodule symbiosis in the common bean.