Background: Triacylglycerols (TAGs) are the main composition of plant seed oil. Long-chain acyl-coenzyme A synthetases (LACSs) catalyze the synthesis of long-chain acyl-coenzyme A, which is one of the primary substrates for TAG synthesis. In Arabidopsis, the LACS gene family contains nine members, among which LACS1 and LACS9 have overlapping functions in TAG biosynthesis. However, functional characterization of LACS proteins in rapeseed have been rarely reported. Results: An orthologue of the Arabidopsis LACS2 gene (BnLACS2) that is highly expressed in developing seeds was identified in rapeseed (Brassica napus). The BnLACS2-GFP fusion protein was mainly localized to the endoplasmic reticulum, where TAG biosynthesis occurs. Interestingly, overexpression of the BnLACS2 gene resulted in significantly higher oil contents in transgenic rapeseed plants compared to wild type, while BnLACS2-RNAi transgenic rapeseed plants had decreased oil contents. Furthermore, quantitative real-time PCR expression data revealed that the expression of several genes involved in glycolysis, as well as fatty acid (FA) and lipid biosynthesis, was also affected in transgenic plants. Conclusions: A long chain acyl-CoA synthetase, BnLACS2, located in the endoplasmic reticulum was identified in B. napus. Overexpression of BnLACS2 in yeast and rapeseed could increase oil content, while BnLACS2-RNAi transgenic rapeseed plants exhibited decreased oil content. Furthermore, BnLACS2 transcription increased the expression of genes involved in glycolysis, and FA and lipid synthesis in developing seeds. These results suggested that BnLACS2 is an important factor for seed oil production in B. napus.

The floral-dip method of transformation by immersion of inflorescences in a suspension of Agrobacterium was applied in the Brassica napus transformation, but it involves a number of relatively time-consuming and laborious steps, including manipulating an Agrobacterium tumefaciens culture and aseptic procedures for the selection of plant lines harboring antibiotic-selection markers. It calls for a long time to prepare the buffered media. To circumvent these bottlenecks, we have developed a rapid and simple method. We find that Brassica napus can be transformed by dipping directly into an Agrobacterium tumefaciens culture supplemented with surfactant, eliminating the need for media exchange to a buffered solution. We report a method of transformant selection by soaking seeds with antibiotic. These methods save time and money, and reduce the possibility of contamination.

ABSTRACT:Effects of different forms of dietary selenium (Se) supplementation on gene expression of cytoplasmic thioredoxin reductase (TrxR1), selenoprotein P (SelP), and selenoprotein W (SelW) in broilers were investigated. A total of six hundred Ross 308 broilers (1-day-old) with similar body weight were randomly divided into three groups, each of which included 5 replicates of 40 birds. These three treatments received the same basal diet with only background Se level of 0.04 mg Se/kg, supplemented with 0.15 mg Se/kg as sodium selenite (SS) or l-selenomethionine (l-Se-Met) or d-selenomethionine (d-Se-Met) for 42 days. The SS supplemented diet increased TrxR1 activity in liver (P < 0.01) and kidney (P < 0.01) as well as SelP concentration in serum (P < 0.05) and liver (P < 0.01) more than the d-Se-Met supplemented diet. The addition of SS also highly increased liver (P < 0.01) and kidney (P < 0.01) TrxR1 activities of broilers in comparison with broilers fed l-Se-Met diet. In addition, liver TrxR1 activity in l-Se-Met group was higher than that in d-Se-Met group (P < 0.05). Liver and kidney mRNA levels of TrxR1 and SelP as well as breast muscle SelW mRNA level were significantly increased by l-and d-Se-Met supplementation in comparison with SS supplementation (P < 0.01), while the d-Se-Met group showed more effective (P < 0.01) than the l-Se-Met group in increasing the mRNA levels of TrxR1 and SelP in liver and kidney. Therefore, dietary l-Se-Met and d-Se-Met supplementation could improve mRNA levels of different selenoproteins studied and reduce amounts of TrxR1 and SelP in broilers compared with SS. Besides, l-Se-Met is more effective than d-Se-Met in raising TrxR1 activity and decreasing mRNA abundance of TrxR1 and SelP in broilers.

Long chain acyl-coenzyme A synthetases (LACSs) activate free fatty acids to acyl-CoA thioesters, and play important roles in the biosynthesis and degradation of lipids. In this study, a Glycine max(L.) LACS gene, designated as GmLACS, has been isolated through in silicon cloning. The gene is 2,219 bp with an open reading frame (ORF) of 1,989 bp, which encodes a LACS with 662 amino acid residues, with the isoelectric point of 6.42 and the calculated molecular mass of 65.6 kDa. Sequence analysis showed that GmLACS possessed typical domains of LACSs. Real-time quantitative PCR data analysis suggested that GmLACS was hightly expressed in leaves and young pods.

Long chain acyl-coenzyme A synthetases (LACSs) activate free fatty acid to acyl-CoA thioesters, and play important roles in the biosynthesis and degradation of lipids. In this study, four cDNAs (Complementary DNA) encode long chain fatty acyl-CoA synthetase activity has been found in Brassica napus. Sequence analysis indicated that the four LACSs possessed typical molecular characteristics of LACS. Compared with low oil content varieties seed, the four genes are strongly expressed in high oil content varieties seeds at 35 days after pollination (DAP). The expression pattern suggested that the four LACSs might be involved in the biosynthesis of lipids and oil accumulation in rapeseed.