RESUMEN
The fatty acid component of commodity soybean seeds typically consists of approximately 12-15% saturated fatty acids in the form of palmitic acid and stearic acid. An important goal in soybean breeding is the reduction of saturated fats, in order to produce healthier vegetable oils for food applications. Genetic approaches have been instrumental in reducing levels of palmitic acid, which is the most abundant saturated fat in soybean seeds. In this study we describe a new mutant allele of the FATB1a gene that encodes a palmitoyl-acyl carrier protein thioesterase. The mutation is expected to result in early termination of the FATB1A protein and mutant seeds carrying this allele contain 5.5% palmitic acid. This new allele can be introduced into conventional soybean lines, alone or in combination with other modifications to generate soybean lines with improved oil composition.
Asunto(s)
Glycine max , Ácido Palmítico , Ácidos Grasos/metabolismo , Mutación , Ácido Palmítico/metabolismo , Fitomejoramiento , Aceites de Plantas/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Semillas/genética , Semillas/metabolismo , Aceite de Soja/metabolismo , Glycine max/genética , Glycine max/metabolismoRESUMEN
Soybean seeds produce valuable protein that is a major component of livestock feed. However, soybean seeds also contain the anti-nutritional raffinose family oligosaccharides (RFOs) raffinose and stachyose, which are not digestible by non-ruminant animals. This requires the proportion of soybean meal in the feed to be limited, or risk affecting animal growth rate or overall health. While reducing RFOs in soybean seed has been a goal of soybean breeding, efforts are constrained by low genetic variability for carbohydrate traits and the difficulty in identifying these within the soybean germplasm. We used reverse genetics Targeting Induced Local Lesions in Genomes (TILLING)-by-sequencing approach to identify a damaging polymorphism that results in a missense mutation in a conserved region of the RAFFINOSE SYNTHASE3 gene. We demonstrate that this mutation, when combined as a double mutant with a previously characterized mutation in the RAFFINOSE SYNTHASE2 gene, eliminates nearly 90% of the RFOs in soybean seed as a proportion of the total seeds carbohydrates, and results in increased levels of sucrose. This represents a proof of concept for TILLING by sequencing in soybean.
Asunto(s)
Alelos , Galactosiltransferasas , Glycine max/genética , Polimorfismo Genético , Análisis de Secuencia de ADN , Galactosiltransferasas/genética , Galactosiltransferasas/metabolismo , Genética de Población , Oligosacáridos/genética , Oligosacáridos/metabolismo , Fitomejoramiento , Rafinosa/genética , Rafinosa/metabolismo , Semillas/genética , Semillas/metabolismo , Proteínas de Soja/genética , Proteínas de Soja/metabolismo , Glycine max/metabolismoRESUMEN
Soybean oil has a wide variety of uses, and stearic acid, which is a relatively minor component of soybean oil is increasingly desired for both industrial and food applications. New soybean mutants containing high levels of the saturated fatty acid stearate in seeds were recently identified from a chemically mutagenized population. Six mutants ranged in stearate content from 6-14% stearic acid, which is 1.5 to 3 times the levels contained in wild-type seed of the Williams 82 cultivar. Candidate gene sequencing revealed that all of these lines carried amino acid substitutions in the gene encoding the delta-9-stearoyl-acyl-carrier protein desaturase enzyme (SACPD-C) required for the conversion of stearic acid to oleic acid. Five of these missense mutations were in highly conserved residues clustered around the predicted di-iron center of the SACPD-C enzyme. Co-segregation analysis demonstrated a positive association of the elevated stearate trait with the SACPD-C mutation for three populations. These missense mutations may provide additional alleles that may be used in the development of new soybean cultivars with increased levels of stearic acid.
Asunto(s)
Glycine max/genética , Glycine max/metabolismo , Oxigenasas de Función Mixta/genética , Mutación , Semillas/genética , Semillas/metabolismo , Ácidos Esteáricos/metabolismo , Secuencia de Aminoácidos , Ácidos Grasos/metabolismo , Genotipo , Oxigenasas de Función Mixta/química , Datos de Secuencia Molecular , Fenotipo , Alineación de SecuenciaRESUMEN
Sixteen replicate microcosms were inoculated with a mixed assemblage of heterotrophic bacteria and provided with discrete pulses of protein as carbon and energy source. The dynamics of community structure were monitored by 16S rRNA gene polymerase chain reaction denaturant gradient gel electrophoresis (PCR-DGGE). The results were consistent with a strong role for biological interactions in maintaining diversity. Replicate microcosms developed different microbial communities. For systems exposed to nutrient pulses every 7 days, the number of DGGE bands averaged 13 +/- 4 (mean +/- SD) and the Dice similarity coefficient between pairs ranged from 0.08 to 0.67. In each of 16 systems provided protein once each day, there were dynamic changes over the first 30 days but community composition was stable over the next 20 days. However, most systems differed from each other; two-thirds of the pairwise comparisons had similarity coefficients in the range of 0.35-0.63. These 16 systems contained 10 +/- 2 phylotypes (mean +/- SD) and in aggregate 34 phylotypes were found in the 16 systems. Most phylotypes were found in < 25% of the systems, and there were not strong networks of association among phylotypes.
Asunto(s)
Bacterias/metabolismo , Fenómenos Fisiológicos Bacterianos , Ecosistema , Periodicidad , Bacterias/clasificación , Bacterias/genética , Bacterias/crecimiento & desarrollo , Medios de Cultivo/química , Electroforesis en Gel de Poliacrilamida/métodos , Cinética , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADNRESUMEN
In nature, microbes are subject to nutrient fluxes. As the periodicity of nutrient flux lengthens, different physiological traits may be selected. The competitive exclusion principle stipulates that one organism will dominate these systems; however, interspecies interactions may produce a dynamic microbial community. These issues were investigated in chemostats pulsed with gelatin. Chemostats were run over 30 days with substrate addition continuously or at intervals of 0.5, 1 or 3 days. Growth rates were similar between pulse intervals. Ectoaminopeptidase activity levels remained relatively constant within a pulse interval. Bacterial community structure was monitored using denaturing gradient gel electrophoresis of PCR products of the 16S rRNA gene. There were dynamic changes at all periodicities; however, the pace of these changes decreased over time. Final communities were not identical between different treatments. The structure of persistent vs. active microbial populations was compared by denaturing gradient gel electrophoresis of the PCR and reverse transcriptase-PCR amplicons of 16S rDNA and rRNA templates, respectively. For all the chemostats, the rRNA profiles were not identical to the rDNA profiles for a sample. These experiments demonstrate that complex community dynamics can occur under environmental heterogeneities that are modest relative to those found in natural aquatic habitats. Furthermore, the physiological functionality of these dynamic communities was stable.
Asunto(s)
Aminopeptidasas/metabolismo , Bacterias/crecimiento & desarrollo , Medios de Cultivo , Ecosistema , Periodicidad , ARN Ribosómico 16S/química , Aguas del Alcantarillado/microbiología , Biomasa , Reactores Biológicos/microbiología , Electroforesis/métodos , CinéticaRESUMEN
When microbes are subjected to temporal changes in nutrient availability, growth rate and substrate affinity can contribute to competitive fitness and thereby affect microbial community structure. This hypothesis was tested using planktonic bacterial communities exposed to nutrient additions at 1-, 3-, 7-, or 14-day intervals. Growth rates after nutrient addition were inversely proportional to the pulse interval and declined from 0.5 h(-1) to 0.15 h(-1) as the pulse interval increased from 1 to 14 days. The dynamics of community structure were monitored by 16S rRNA gene PCR-denaturing gradient gel electrophoresis. At pulse intervals of more than 1 day, the community composition continued to change over 130 days. Although replicate systems exposed to the same pulse interval were physiologically similar, their community compositions could exhibit as much dissimilarity (Dice similarity coefficients of <0.5) as did systems operated at different intervals. Bacteria were cultivated from the systems to determine if the physiological characteristics of individual members were consistent with the measured performance of the systems. The isolates fell into three bacterial divisions, Bacteroidetes, Proteobacteria, and Actinobacteria. In agreement with community results, bacteria isolated from systems pulsed every day with nutrients had higher growth rates and ectoaminopeptidase specific activities than isolates from systems pulsed every 14 days. However, the latter isolates did not survive starvation longer than those provided with nutrients every day. The present study demonstrates the dynamic nature of microbial communities exposed to even simple and regular environmental discontinuities when a substantial pool of species that can catabolize the limiting substrate is present.