| Brassica napus is a major oil-producing crop, which is an important source of the edible oil supply and widely used in industry and bioenergy. Because B. napus seeds contain more beneficial unsaturated fatty acids, high protein and other advantages, it is becoming more and more popular in food research in recent years. However, many restrictive factors are serious threat to rapeseed production in China. Sclerotinia. sclerotiorum is the main limiting factor to the rapeseed production. In addition to exogenous diseases and insect pests, the endogenous factors, such as seed germination rate, oil content and so on, also seriously affect the yield of rape. It is of great significance to improve the yield of rape. In addition to the traditional breeding, in recent years, more and more researchers focus on transgenic technology to improve B. napus crops.GDSL lipases/esterases are super lipase families which play roles in abiotic stress, pathogen defense, seed development and lipid metabolism.In this paper, we mainly studied the biological functions of two lipase and esterase genes, exogenous Arabidopsis thaliana AtGDSL1 and endogenous B. napus BnGDSL1. Bioinformatics analysis showed that they have five conserved domains and 85.99% amino acid sequence similarity. We also found that they have a significant impact on the resistance to S. sclerotiorum, seed germination rate and oil content. Bioinformatics analysis also found that AtGDSL1 and BnGDSL1 are hydrophilic proteins, containing a 25 amino acid trans-membrane domain in the N terminal, which may be conducive to subsequent soluble protein purification. These structure characteristics suggest that they may have similar functions.Yan Suzhen in our lab had proved BnGDSL1 was a lipase. To further research the lipase activity of AtGDSL1, we constructed the AtGDSL1 prokaryotic expression recombinant vector. AtGDSL1 recombinant protein was excpressed and purified in E. coli, then used for lipase activity determination. Lipase activity increased from one hour to five hours and the highest lipase activity was observed at five hours with 7.2 mM/mg activity.To further study the function of AtGDSL1 and BnGDSL1 in oilseed rape, AtGDSL1 and BnGDSL1 overexpression plants as well as BnGDSL1 suppression plants were constructed. First of all, through inoculation the transgenic plants’ leaf with S. sclerotiorum, we found that overexpression of AtGDSL1 plants significantly improved S. sclerotiorum resistance. However, BnGDSL1 has no significant effect on S. sclerotiorum. Secondly, we found that the seed germination rate of AtGDSL1 and BnGDSL1 overexpression plants were significantly increased, while the seed germination rate was inhibited in BnGDSL1 suppression plants. Finally, by near infrared and nuclear magnetic resonance(NMR) found the seed oil content of BnGDSL1 suppression plants were increased, while AtGDSL1 and BnGDSL1 overexpression plants decreased slightly.Our study showed that S. sclerotiorum resistance, seed germination rate and oil content were improved by regulating the GDSL lipase transcription level. This study was helpful to understand the molecular mechanism of plant disease resistance and provided clues for the prevention of S. sclerotiorum. In rapeseed production, the increase of seed germination rate and oil content were of great significance in breeding practice. |
