Studies On The Effects Of Plant Growth Regulators On Glucosinolates And The Expression Of Related Genes In Pakchoi

Glucosinolates (β-thioglucoside N-hydroxysulfates, GSs) are nitrogen-and sulfur-containing anionic natural products, one of the important secondary metabolites found in Brassicaceae plants. Glucosinolates have been proved to have strong relations with Brassica vegetable’s flavors, tastes, nutrition and plant defense system. Glucosinolates also have beneficial effects on human health. The GSs in plant are mostly determined by plant genotypes, as well as affected by plant growth conditions, environment factors and plant growth regulators. Modifying the GS profiles and concentrations by genetic engineering and other factors, to get more beneficial GSs for human health, or to promote the capacity of plant defense to environment stress, has become a hot research topic in recent years. For the development of good quality vegetable crops and sustainable agriculture, it is of great importance to study the effects of plant growth regulator on plant GS metabolism.This study uses the traditional and important Chinese cruciferous vegetable pak choi (Brassica rapa L. ssp. Chinensis var. communis) to study the effects of brassinosteroid (BR), polyamine (PA), insecticide, herbicide, nitric dioxide (NO), hydrogen sulfide (H2S) and sulfur dioxide (SO2) on plant GS accumulation. We also isolated the GSs biosynthetic gene BcCYP83A1and BcCYP83B1from pak choi and analyzed the gene expression pattern in leaves of seven pak choi cultivars and in three organs of pak choi. By analyzing the GS accumulation and gene expression, we explicated the effects and mechanisms of plant regulators on plant GSs synthesis. The following are the major results:(1) The effects of exogenous BR and PA on plant growth and GS contents were studied in pak choi plants grown in nutrient solution. The results show BR increased the shoot aliphatic and total GSs contents, decreased the indole GS contents of pak choi. The increased aliphatic GSs by BR may due to the enhanced amino acid and glucose accumulation. And the decreasing of indole GSs by BR may have involvements with auxin metabolism and plant defense system.0.1mM supermine (spm) and1.0mM supermidine (spd) both increased the aliphatic GSs, aromatic GSs and total GSs. The effects of exogenous PA on GS contents of pak choi may involve in the inner PA and ethylene metabolisom.(2) The effects of four insecticides and five herbicides on plant growth and GS contents were investigated in pak choi plants grown in nutrient solution. All the insecticides increased the shoot aliphatic GSs, aromatic GSs, indole GSs and total GSs of pak choi. It is speculated that the increasing GS contents were the strategy of plant response to insecticide stress. All herbicides decreased the aliphatic GS contents in shoots of pak choi, and these may due to the decreasing effects of herbicides on aliphatic amino acid. Besides of the decreasing of aliphatic GSs, quizalofop-p-ethyl also increased the aromatic GSs and indole GSs contents which leading to the increasing of total GSs. The influences of herbicides on pak choi GS contents were short term effects.(3) The effects of exogenous NO, H2S and SO2on plant growth and GS contents were studied in pak choi plants grown in nutrient solution. The results show proper concentration of exogenous NO have short term (5day) positive effects on GS accumulation in shoots of pak choi, but have negative effects on GS contents in long term (10day). Exogenous NO also affected the profile and contents of GSs in roots of pak choi. Both H2S and Na2SO4treatments increased the aliphatic GSs, aromatic GSs, indole GSs and total GSs in shoots of pak choi. The GS contents were increased followed by the increasing of H2S concentrations. However, the effects of different concentration Na2SO4treatments on GSs had no difference. The decreased GS contents treated by1.0mM and10mM NaHSO3show SO2can not be used as sulfur source by pak choi, but have negative effects on plant growth and GS accumulation. The positive effects of H2S on GSs accumulation were short term effects.(4) The aliphatic and indole GS biosynthetic gene BcCYP83Al and BcCYP83B1were isolated from pak choi. Two copies of BcCYP83A1, named BcCYP83A1-1and BcCYP83A1-2and one copy of BcCYP83B1were isolated. The predicted amino acid sequences of BcCYP83Al-1, BcCYP83A1-2and BcCYP83Bl shared high sequence identity of87.65,86.48and95.59%to the corresponding ones in Arabidopsis, and98.80,98.61and98.80%to the corresponding ones in Brassica pekinensis (Chinese cabbage), respectively. The cloning of the GS biosynthetic genes would have beneficial to modify the GS profiles and contents using genetic methods.(5) Quantitative real-time PCR (qRT-PCR) were used to analysis the expression levels of GS biosynthetic genes in different cultivars. different organs and as affected by plant growth regulators. The results indicated that both BcCYP83A1and BcCYP83B1expressed in roots, leaves and petioles of pak choi, while the transcript abundances of BcCYP83A1were higher in leaves than in petioles and roots, whereas BcCYP83B1 showed higher abundances in roots. The expression levels of GS biosynthetic genes were consistent with the GS profile accumulation in leaves of seven cultivars and three organs. All the examined plant growth regulators affected the GS biosynthetic genes expression and the expression levels of GS biosynthetic genes were consistent with the GS profile accumulation as affected by plant growth regulators.