RESUMEN
About 99% of soil microorganisms are unculturable. However, advances in molecular biology techniques allow for the analysis of living microorganisms. With the advent of new technologies and the optimization of previous methods, various approaches to studying gene expression are expanding the field of microbiology and molecular biology. Methods used for RNA extraction, DNA microarrays, real-time PCR, competitive RT-PCR, stable isotope probing and the use of reporter genes provide methods for detecting and quantifying gene expression. Through the use of these methods, researchers can study the influence of soil environmental factors such as nutrients, oxygen status, pH, pollutants, agro-chemicals, moisture and temperature on gene expression and some of the mechanisms involved in the responses of cells to their environment. This review will also address information gaps in bacterial gene expression in soil and possible future research to develop an understanding of microbial activities in soil environments.
Asunto(s)
Bacterias/genética , Hongos/genética , Técnicas Microbiológicas/métodos , Microbiología del Suelo , Suelo/análisis , Bacterias/aislamiento & purificación , ADN Complementario/análisis , Hongos/aislamiento & purificación , Expresión Génica , Regulación de la Expresión Génica , Genes Reporteros , Concentración de Iones de Hidrógeno , Marcaje Isotópico , Análisis de Secuencia por Matrices de Oligonucleótidos , Oxígeno/metabolismo , Reacción en Cadena de la Polimerasa , ARN Bacteriano/análisis , ARN Bacteriano/genética , ARN de Hongos/análisis , ARN de Hongos/genética , Contaminantes del Suelo/metabolismo , TemperaturaRESUMEN
Roundup Ready (RR) soybeans containing recombinant Agrobacterium spp. CP4 5-enol-pyruvyl-shikimate-3-phosphate synthase (cp4 epsps) genes tolerant to the herbicide glyphosate are extensively grown worldwide. The concentration of recombinant DNA from RR soybeans in soil aggregates was studied due to the possibility of genetic transformation of soil bacteria. This study used real-time PCR to examine the concentration of cp4 epsps in four field soil aggregate size classes (>2000 microm, 2000-500 microm, 500-250 microm and <250 microm). Aggregates over 2000 microm in diameter had significantly greater gene concentrations than those with diameters under 2000 microm. The >2000 mum fraction contained between 66.62% and 99.18% of total gene copies, although it only accounted for about 30.00% of the sampled soil. Aggregate formation may facilitate persistence of recombinant DNA.
Asunto(s)
3-Fosfoshikimato 1-Carboxiviniltransferasa/análisis , Proteínas Bacterianas/análisis , ADN de Plantas/análisis , Glycine max/química , Plantas Modificadas Genéticamente/química , Suelo/análisis , 3-Fosfoshikimato 1-Carboxiviniltransferasa/genética , Proteínas Bacterianas/genética , ADN de Plantas/genética , Lectinas/análisis , Lectinas/genética , Tamaño de la Partícula , Proteínas de Plantas/análisis , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente/genética , Rhizobium/enzimología , Glycine max/genéticaRESUMEN
Glyphosate-tolerant, Roundup Ready (RR) soybeans account for about 57% of all genetically modified (GM) crops grown worldwide. The entry of recombinant DNA into soil from GM crops has been identified as an environmental concern due to the possibility of their horizontal transfer to soil microorganisms. RR soybeans contain recombinant gene sequences that can be differentiated from wild-type plant and microbial genes in soil by using a sequence-specific molecular beacon and real-time polymerase chain reaction (PCR). A molecular beacon-based real-time PCR system to quantify a wild-type soybean lectin ( le1) gene was designed to compare amounts of endogenous soybean genes to recombinant DNA in soil. Microcosm studies were carried out to develop methodologies for the detection of recombinant DNA from RR soybeans in soil. RR soybean leaf litterbags were imbedded in the soil under controlled environmental conditions (60% water holding capacity, 10/15 degrees C, and 8/16 h day/night) for 30 days. The soybean biomass decomposition was described using a single-phase exponential equation, and the DNA concentration in planta and in soil was quantified using real-time PCR using sequence-specific molecular beacons for the recombinant cp4 epsps and endogenous soybean lectin ( le1) genes. The biomass of RR soybean leaves was 8.6% less than nontransgenic (NT) soybean leaves after 30 days. The pooled half-disappearance time for cp4 epsps and le1 in RR and of le1 in NT soybean leaves was 1.4 days. All genes from leaves were detected in soil after 30 days. This study provides a methodology for monitoring the entry of RR and NT soybean DNA into soil from decomposing plant residues.
Asunto(s)
ADN Recombinante/análisis , Glycine max/genética , Hojas de la Planta/genética , Plantas Modificadas Genéticamente/genética , Reacción en Cadena de la Polimerasa , Suelo/análisis , ADN de Plantas/análisis , Tolerancia a Medicamentos/genética , Glicina/análogos & derivados , Lectinas de Plantas/genética , Proteínas de Soja/genética , GlifosatoRESUMEN
We grew plants of nine soybean varieties, six of which were genetically modified to express transgenic cp4-epsps, in the presence of Bradyrhizobium japonicum and arbuscular mycorrhizal fungi. Mycorrhizal colonization and nodule abundance and mass differed among soybean varieties; however, in no case was variation significantly associated with the genetic modification.