RESUMEN
KEY MESSAGE: Proteomic and lipidomics analyses of WT and GmDGAT1-2 transgenic soybeans showed that GmDGAT1-2 over-expression induced lipoxygenase down-regulatation and oleoin up-regulatation, which significantly changed the compositions and total fatty acid. The main goal of soybean breeding is to increase the oil content. Diacylglycerol acyltransferase (DGAT) is a key rate-limiting enzyme in fatty acid metabolism and may regulate oil content. Herein, 10 GmDGAT genes were isolated from soybean and transferred into wild-type (WT) Arabidopsis. The total fatty acid was 1.2 times higher in T3 GmDGAT1-2 transgenic Arabidopsis seeds than in WT. Therefore, GmDGAT1-2 was transferred into WT soybean (JACK), and four T3 transgenic soybean lines were obtained. The results of high-performance gas chromatography and Soxhlet extractor showed that, compared with those of JACK, oleic acid (18:1), and total fatty acid levels in transgenic soybean plants were much higher, but linoleic acid (18:2) was lower than WT. Palmitic acid (16:0), stearic acid (18:0), and linolenic acid (18:3) were not significantly different. For mechanistic studies, 436 differentially expressed proteins (DEPs) and 180 differentially expressed metabolites (DEMs) were identified between WT (JACK) and transgenic soybean pods using proteomic and lipidomics analyses. Four lipoxygenase proteins were down-regulated in linoleic acid metabolism while four oleosin proteins were up-regulated in the final oil formation. The results showed an increase in the total fatty acid and 18:1 composition, and a decrease in the 18:2 composition of fatty acid. Our study brings new insights into soybean genetic transformation and the deep study of molecular mechanism that changes the total fatty acid, 18:1, and 18:2 compositions in GmDGAT1-2 transgenic soybean.
Asunto(s)
Diacilglicerol O-Acetiltransferasa/genética , Glycine max/genética , Lipooxigenasa/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas de Plantas/metabolismo , Aceite de Soja/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Diacilglicerol O-Acetiltransferasa/metabolismo , Regulación de la Expresión Génica de las Plantas , Ácido Linoleico/genética , Ácido Linoleico/metabolismo , Lipidómica/métodos , Lipooxigenasa/genética , Proteínas de la Membrana/genética , Familia de Multigenes , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente , Proteómica/métodos , Semillas/genética , Semillas/metabolismo , Aceite de Soja/genética , Proteínas de Soja/genética , Proteínas de Soja/metabolismo , Glycine max/metabolismoRESUMEN
This study was aimed to optimize the extraction (CO2 supercritical extraction) process and purification (ethanol washing and decontaminating) process of total fatty acid in Brassica campestris pollen. With the extraction yield of total fatty acid as index, CO2 supercritical extraction of total fatty acid in B. campestris polle was optimized by central composite design-response surface methodology. Ethanol washing and decontaminating was done for the extract through orthogonal design with content of total fatty acid (linolenic acid amide, linolenic acid glyceride, linolenic acid and palmitic acid) as the indexes. The optimum parameters of CO2 supercritical extraction technology were as follows: extraction pressure of 35 MPa, extraction temperature of 60 âand extraction time of 3 h. When extract of supercritical fluid was purified by 50 times of 80% ethanol for 1.5 h, the content of total fatty acid can reach 60%. In addition, this process was stable and steady, provided reliable basis for production.