镉胁迫对小白菜(Brassica campestris L.)抗氧化机理的影响

采用水培的方法,研究了不同Cd2+水平(0、1、2.5、5、10、15mg·L-1)对小白菜(Brassica campestris L.)抗氧化酶(SOD、POD、CAT)活性、非酶物质(SH、GSH、PCs)含量以及生长的影响。结果表明,小白菜叶片和根系SOD活性随Cd处理质量浓度的增加呈降低的变化趋势,POD、CAT活性以及MDA含量随Cd处理质量浓度的增加而增加。小白菜的生物量、根长、株高、叶绿素质量分数随Cd处理质量浓度的增加显著降低(P<0.05),表明Cd抑制了小白菜的生长,破坏了叶绿素的合成。小白菜地上、地下部镉质量分数均随Cd处理质量浓度的增加而显著增加(P<0.05)。当Cd处理质量浓度为15mg·L-1,小白菜地上、地下部镉质量分数分别达到637.5、1663.0mg·kg-1,表明小白菜对Cd有良好的富集效果。小白菜根系与叶片中SH、GSH和PCs含量均随Cd处理质量浓度增加而增加的变化趋势,表明SH、GSH和PCs在解毒小白菜Cd毒害中起着重要作用。

Role of nitrification inhibitor DMPP(3,4-dimethylpyrazole phosphate) in NO^-_3-N accumulation in greengrocery( Brassica campestris L. ssp. chinensis ) and vegetable soil

The influence of nitrification inhibitor(NI) 3,4 dimethylpyrazole phosphate(DMPP) on nitrate accumulation in greengrocery( Brassica campestris L. ssp. chinensis ) and vegetable soil at surface layer were investigated in field experiments in 2002 and 2003 Results showed that NI DMPP took no significant effect on yields of edible parts of greengrocery, but it could significantly decrease NO - 3 N concentration in greengrocery and in vegetable soil at surface layer. In addition, NI DMPP could reduce the NO - 3 N concentration during the prophase stage of storage.

Effect of Nitrogen and Sulfur Supply on Glucosinolates in Brassica campestris ssp.chinensis

Glucosinolates （GSs） are a group of plant secondary metabolites containing abundant nitrogen （N） and sulfur （S） mainly in Brassica and have the beneficial effects on human health including anti-carcinogenic, cholesterol-reducing and other pharmacological effects. The objective of this study was to investigate the effect of different concentrations of N （5, 10, and 20 mmol L-a, denoted by N5, N10 and N20） and S （0,5, 1, and 2 mmol L^-1, denoted by S0.5, S1 and S2） on the yield and GSs in pakchoi （Brassica campestris L. ssp. chinensis var. communis） in hydroponics. Results showed that N10 and N20 significantly enhanced the yield compared with N5, however, N20 had a negative effect relative to N10. Only with N10 and N20 low S supply （S0.5） reduced the yield. The concentrations of aliphatic GSs, aromatic GS and total GSs were enhanced by N5 and indolyl GSs were enhanced by N20. S2 enhanced the concentration of individual GS and total GSs. The concentrations of indolyl GSs were maximized in N20S2 treatment, whereas the highest concentrations of aliphatic GSs, aromatic GS and total GSs were found in N5S2 treatment. Effects of N and S on aliphatic GSs were higher than on indolyl GSs. The results suggest that the accumulation of aliphatic GSs and aromatic GS could be enhanced by low N and high S and restricted by high N while that of indolyl GSs could be enhanced by high N and high S.

A Genetic Linkage Map of Brassica campestris L.ssp. pekinensis (syn. B. rapa L. ssp. pekinensis)

A molecular genetic map of Chinese cabbage was constructed with a 102 recombinant inbred (RI) population from a cross of two cultivated Chinese cabbage lines 177 and 276, using AFLP and RAPD markers. 352 markers including 265 AFLP markers and 87 RAPD markers were integrated into 17 linkage groups. It covered a total of 2 665. 7 cM with an average interval of 7. 6 cM. AFLP marker is efficient for map construction while it easily forms clusters to cause big gaps in map. A total of 13.92 % abnormal segregation markers distributed in the map. The molecular genetic map is fundamental for gene localization, comparative genomics, and QTL mapping of important agronomic traits.

Differential Expression Analysis of Genic Male Sterility A／B Lines in Chinese Cabbage—Pak—Choi（Brassica campestris ssp.chinensis Makino）

To determine differential expression of genie male sterility A/B lines in Chinese cabbage-pak-choi (Brassica campestris ssp. chinensis Makino var. communis Tsen et Lee), we used the RNA fingerprinting technique, cDNA-AFLP analysis, in different developmental stages and different tissues. While no obvious differential expressions were observed in rosette leaves, florescence leaves, and scapes, some differential expressions were found in alabstrums of A/B lines and among leaves, scapes and alabstrums. We analyzed the al-abstrums collected in different developmental stages with 10 primer combinations. We got a unique band between middle size alabstrums and large alabstrums in B line in one of the ten pair primers, and in another one pair, one band reflecting a higher gene-expression level in A line than that in B line was obtained. No unique bands were found with the other primer combinations. The bands reflecting different gene-expression level were confirmed by Northern hybridization. The results indicated that cDNA-AFLP was a suitable tool for studying differential expression of genie male sterility in plants. SDS-polyacrylamide gel electrophoresis patterns of soluble proteins further verified the difference in A/B lines.

Nitrogen Dioxide-Induced Responses in Brassica campestris Seedlings: The Role of Hydrogen Peroxide in the Modulation of Antioxidative Level and Induced Resistance

This article investigates the responses of Brassica campestris seedlings to an acute level of nitrogen dioxide （NO2） exposure in a plant growth chamber, and examines whether pretreating plants with hydrogen peroxide （H2O2） will alleviate NO2-caused injury. Twenty-eight-day-old B. campestris plants sprayed with 10 mmol L^-1 H2O2 aqueous solution （corresponding to approximate 1.0 mg H2O2 per single plant） were exposed to different concentrations of NO2 （0.25, 0.5, 1.0, and 2.0 μL L^-1, respectively） for 24 h under controlled environment. To measure the plant biomass, the plants were fumigated with the same NO2 concentrations as mentioned above for 7 h per day （8.00-15.00） for 7 days. As a control, charcoal filtered air alone was applied. Data were collected on plant biomass, total chlorophyll, photosynthetic rate, stomatal conductance, nitrate and nitrate reductase （NR）, antioxidative enzymes, ascorbate （ASA）, and malondialdehyde （MDA）, immediately after exposure. The results showed that exposure to a moderate dose of NO2 （e.g., 0.25 μL L^-1） had a favorable effect on plants, and the dry weight of the above-ground part increased, whereas the exposure to high NO2 concentrations （e.g., 0.5 μL L^-1 or higher） caused a reduction in the plant biomass and the total chlorophyll, when compared with the control. In addition, at 0.5 μL L^-1 or higher NO2 concentrations, prominent increases in the MDA level and superoxide dismutase （SOD） and NR activities were observed. Exposure to 1 μL L^-1 and higher NO2 resulted in necroses appearing on older leaves, and an increase in catalase （CAT） activity, decrease in ASA content, increased accumulation of NO3^-, and reduction in photosynthesis, when compared with the controls. No changes were detected in stomatal conductance under NO2 fumigation. The pretreatment with 10 mmol L^-1 H2O2 alleviated significantly NO2- caused biomass decrease and photosynthetic inhibition when compared with H2O2-untreated plants. Under NO2 fumigation, further induction in SOD and CAT activities occurred in H2O2 treated plants when compared with H2O2- untreated plants. The effect of NO2 on the ASA and MDA contents was also absent in H2O2-treated plants. However, the H2O2 treatment did not alter the nitrate content and NR activity in plants under NO2 fumigation. The H2O2 treatment caused a lower rate of stomatal conductance. Taken together, these data suggest that fumigation with an acute level of NO2 causes oxidative damage to B. campestris seedlings. The H2O2 pretreatment markedly protects plants against NO2 stress and this may be associated with inducible antioxidative level. NO2 fumigation contributes, at least in part, to the enhanced levels of nitrate in B. campestris leaves.

Genetic linkage map of Brassica campestris L.using AFLP and RAPD markers

A genetic linkage map comprised of 131 loci was constructed with an F2 population derived from an inter-subspecific cross between Brassica campestris L.ssp.chinensis cv.aijiaohang” and ssp.rapifera cv.,”'isihai”.The genetic map included 93 RAPD loci,36 AFLP loci and 2 morphological loci organized into 10 main linkage groups(LGs) and 2 small groups,covering 1810.9cM with average distance between adjacent markers being approximately 13.8cM.The map is suitable for identification of molecular markers linked to important agronomic traits.QTL analysis,and even for marker-assisted selection in breeding programs of Chinese cabbage and turnip.

Effect of the Antisense BcMF12 Driven by the BcA9 Promoter on Gene Silencing in Brassica campestris L.ssp.chinensis

The study analyzed the silencing of BcMF12 gene regulated by BcA9 promoter in the transgenic pakchoi and confirmed the effect of antisense BcMF12 gene on the pollen development. A conserved BcMF12 gene fragment was amplified from the cDNA of flower buds in pakchoi （Brassica campestris L. ssp. chinensis, syn. B. rapa L. ssp. chinensis） and was fused to the anther specific BcA9 promoter. The plant antisense expression vector was constructed and then introduced into pakchoi via Agrobacterium-mediated transformation. The transgenic plants were screened by antibiotics and molecular analysis. PCR and Southern blot revealed that the antisense BcMF12-GUS fusion gene regulated by BcA9 promoter was integrated into transgenic plants. Northern blot suggested that the expression of BcMF12 gene was down-regulated significantly. The pollen germination rate of transgenic plants with antisense BcMF12 gene decreased as compared with that of the control plants. The expression of the gene BcMF12 related to the pollen development was inhibited by the antisense BcMF12 driven by BcA9 promoter, which consequently affected the pollen development in pakchoi.

Identifying Cytoplasmic Male Sterile Types of Chinese Cabbage(Brassica campestris L.)by Molecular Markers

Different sterile cytoplasm types of nine cabbage cytoplasmic male sterile materials were identified by molecular marker in the study, in order to better use molecular marker to conduct the assisted breeding in the future. Genomic DNA was isolated from Chinese cabbage by CTAB method. The design of two pairs of specific primers was performed on conserved flanking region of orf138 gene in the GenBank. PCR was performed with genomic DNA of the nine Chinese cabbage materials. The bands were sequenced. The homologous comparison was conducted in NCBI, and finally, the type of sterile cytoplasm was determined. The results showed that the bands were amplified only in four Chinese cabbage male sterile materials with two pairs of specific primers PUPIl and PIII/PIV, while the other five materials did not obtain the relative bands. The result was consistent with the field sterility identification. And then four molecular markers of Chinese cabbage Ogura cytoplasmic male sterility （CMS） were obtained. After conducting a homologous comparative analysis with BLAST in GenBank, it was found that the homologous degree was 100% in specific segments of tbe tbree sterility materials （L1-CI, L3-CI and L3- F1 ） and Ogu orf138 gene （GenBank accession No. ： HQ149728） of the reported broccoli Ogu CMS. The homologous degree of L1-F1 was 99% with a variation point. The type of cytoplasmic male sterility of the other five materials needed further research. Four materials of the nine were identified as the radish cytoplasmic male sterility materials and four molecular markers were obtained.

Genetic Variation and Correlation Analysis on Seed Oil Content and Protein Content of Brassica campestris L.Germplasm Resources

The content of oil and protein in 84 germplasm materials of Brassica campestris L. from all over the country was tested. In Guiyang environment,the oil content and protein content of these materials showed abundant variation,and were divided into three groups. Correlation analysis showed that there was a significant negative correlation between oil content and protein content of B. campestris L. seeds,and the application and formation mechanism of the negative correlation were also discussed.

Effects of Different Cadmium Levels on Active Oxygen Metabolism and H_2O_2-Scavenging System in Brassica campestris L.ssp.chinensis

The effects of different Cd (Cadmium) levels on generation of active oxygen speceies(AOS) and H2O2-scavenging system in the leaves of Brassica campestris L. ssp. chinensiswere studied. The results showed that generation rate, and H2O2 content were enhancedand malondialdehyde (MDA) content increased with the increase of Cd concentrations inthe growth medium. The activities of ascorbate peroxidase (APX), dehydroascorbatereductase (DR) and glutathione reductase (GR) were promoted by the addition of Cd.Exposed to Cd also increased the contents of ascorbate (AsA) and glutathione (GSH) in theleaves.

国五国六柴油车尾气颗粒物对小白菜(Brassica campestris L.)的生态毒性

为研究柴油车尾气颗粒物对植物的毒性效应,本研究以执行国五、国六两种排放标准的柴油车为研究对象,测定其尾气颗粒物中重金属含量,并以小白菜(Brassica campestris L.)为受试植物,分析了两种颗粒物对小白菜的毒性效应.结果表明:(1)国五、国六柴油车尾气颗粒物中13种重金属总含量分别为1.04 g·kg^(-1)和0.47 g·kg^(-1),国五柴油车尾气颗粒物重金属含量高于国六柴油车;(2)在两种颗粒物的高浓度胁迫下,小白菜体内叶绿素以及蛋白质的含量均出现显著降低趋势;(3)随着两种颗粒物浓度的升高,植物体内部分氧化应激的酶指标超氧化物歧化酶(SOD)、过氧化氢酶(CAT)呈先上升后下降的趋势,而过氧化物酶(POD)以及丙二醛(MDA)、谷胱甘肽(GSH)呈上升趋势,颗粒物的污染能对小白菜产生较为显著的氧化胁迫效应.(4)21 d的毒性实验后,国五、国六柴油车尾气颗粒物对小白菜鲜重的半数有效浓度(EC50)值分别为76.27 mg·L^(-1)和104.48 mg·L^(-1).综合来看,国六标准的实施有效控制了颗粒物中重金属的含量,并一定程度地降低了其对小白菜的毒性.

Mechanical Harvesting Effects on Seed Yield Loss,Quality Traits and Profitability of Winter Oilseed Rape (Brassica napus L.)

China is one of the most important rapeseed producing countries in the world. Effective mechanical harvesting time for decreasing harvesting loss of winter oilseed rape has been becoming a critical factor. An elite cultivar Zhongshuang 11 （Brassica napus L.） was employed in two rounds of field experiments from 2009 to 2011. Seeds were sown with machine, three combine harvesting times namely combine harvesting A, B, and C （CHA, CHB, and CHC） were designed and manual harvesting （MH） as control was performed at maturity. The harvesting treatments were determined according to color of pod and seed in the field. Seed yield loss and quality in different treatments were evaluated. Results showed that both seed yields and harvesting losses in 2009-2010 were higher than that in 2010-2011, whereas seed oil contents in 2010-2011 were higher than that in 2009-2010. The highest yield appeared in CHB, which was significantly higher than that in MH. Furthermore, harvesting loss in CHB were 50% that in MH. Seed oil content and chlorophyll exhibited no obvious difference between CHB and MH. Economic profit analysis demonstrated that mechanical sowing/combine harvesting （MS/CH） showed an input/output ratio of 1：1.6, and it was 1：1.2 in mechanical sowing/manual harvesting （MS/MH）. Labor-cost accounted for more than 70% of the total cost in MS/MH, which led to low profitability to a great extent. Our results suggested that CHB was the optimum harvesting time for winter oilseed rape along the Yangtze River.

Effects of harvesting method and date on yield loss and seed quality of rapeseed

China is one of the major rapeseed production countries in the world,but harvesting mechanization was still backward,and high harvest loss was a key inhibiting factor for rapeseed production.To obtain optimum harvesting date for winter rape in the Yangtze River Valley of China,artificial simulated combine harvesting and artificial two-stage harvesting were adopted to find correlations between harvesting date,rape variety,seed oil content,protein content,grain moisture rate,rapeseed straw moisture content,rapeseed unthreshing rate,grain drop loss rate,thousand seeds weight(TSW),and harvesting economic coefficient(HEC).Analysis of variance showed that rapeseed oil content,protein content and TSW were correlated with rape variety;HEC was correlated with harvesting method and rape variety.Rape variety was the dominant factor of rapeseed oil content,protein content,TSW and HEC.Grain moisture rate,un-threshing rate,grain drop loss rate and straw moisture content were correlated with harvesting method and date.Harvesting date was the dominant factor of rapeseed moisture rate,un-threshing rate,grain drop loss rate and straw moisture rate.Single factor tests further proved that harvesting date had less impact on rapeseed oil content,protein content,TSW and HEC.It showed little correlation with rapeseed quality except grain moisture rate.The optimum harvesting date could be known by change in straw moisture content.The combined harvesting should be carried out during the grain moisture content of 15%-20%with TSW stabilized in the highest level.The twostage harvesting cutting should be carried out at the grain moisture content of 35%-40%.This research offered a reference to harvesting method and date for rape cultivated in the Yangtze River Valley.

Effects of Boron Nutrition on ^45Ca Retranslocation and Distribution in Rape(Brassica napus L.)Cultivars

Using tracer techniques with the radioisotope 45Ca, the effects of boron supply from soil and nutrient solution on the retranslocation of 45Ca from leaves to other parts of rape plants were studied. Results indicated that only a small portion of foliar-applied 45Ca could be retranslocated to other parts of the rape plant. There was no pronounced effect of boron level in the soil on 45 Ca retranslocation. Increasing boron concentrations in the nutrient solution significantly reduced 45 Ca radioactivity in root and stem, but increased 45Ca uptake and 45 Ca translocation to the upper leaves of rape plants. It was suggested that45 Ca absorbed by newly developed roots was easily distributed to upper leaves of rape plants. Boron may alter distribution of calcium via its effect on root development or growth of newly developed roots.

浙北地区优质甘蓝型油菜(Brassica napus L.)产量性状与产量的关系

对24个高产优质甘蓝型油菜品种(系)的单株产量及9个产量性状进行统计分析,结果表明,单株有效角果数、每角果实粒数、千粒重对单株产量的正效应达显著或极显著水平;单株有效角果数和每角果实粒数对产量的直接通径系数分别为0.9702和0.8439,且净效应均达极显著正值;而千粒重对产量的直接通径系数虽为正值且较大,但净效应呈负值;结角密度对产量的直接通径系数及净效应均为负值。研究结果分析,浙北优质甘蓝型油菜高产的育种方向,在选择合适结角密度的基础上,重点应放在每角果实粒数和单株有效总角果数两个主要目标性状的选择上,同时兼顾千粒重。

Nutrient deficiency limits population development,yield formation,and nutrient uptake of direct sown winter oilseed rape

Direct-sowing establishment method has great significance in improving winter oilseed rape（Brassica napus L.） production and guaranteeing edible oil security in China. However, nutrient responses on direct sown winter oilseed rape（DOR） performance and population development dynamic are still not well understood. Therefore, five on-farm experiments were conducted in the reaches of the Yangtze River（RYR） to determine the effects of nitrogen（N）, phosphorus（P）, and potassium（K） deficiencies on population density, dry matter production, nutrient uptake, seed yield, and yield components of DOR plants. Four fertilization treatments included the balanced NPK application treatment（NPK, 180 kg N, 39.3 kg P, 100 kg K, and 1.8 kg borax ha–1） and three nutrient deficiency treatments based on the NPK treatment, i.e., –N, –P, and –K. The results indicated that DOR population density declined gradually throughout the growing season, especially at over-wintering and pod-development stages. Nutrient deficiency decreased nutrient concentration in DOR plants, limited dry matter production and nutrient uptake, and thereby exacerbated density reduction during plants growth. The poor individual growth and reduced population density together decreased seed yield in the nutrient deficiency treatment. Averaged across all the experiments, seed yield reduced 61% by N deficiency, 38.3% by P deficiency, and 14.4% by K deficiency. The negative effects of nutrient deficiency on DOR performances followed the order of –N–P–K, and the effects were various among different nutrient deficiencies. Although N deficiency improved DOR emergence, but it seriously limited dry matter production and nutrient uptake, which in turn led to substantial plants death and therefore resulted in a very low harvested density. The P deficiency significantly reduced initial density, limited plants growth, and exacerbated density reduction. The K deficiency mainly decreased individual growth and yield, but did not affect density dynamic. Our results highlighted the importance of balanced NPK application in DOR production, suggesting that management strategy of these nutrients should be comprehensively considered with an aim to build an appropriate population structure with balanced plant density and individual growth.

乌菜BcVIL2基因克隆及春化响应表达分析

【目的】从乌菜(Brassica campestris L.)中克隆VIN3⁃LIKE 2(VIL2)基因并进行生物信息学与表达分析,为进一步研究BcVIL2基因功能奠定基础。【方法】以‘徽乌19’为试验材料,利用同源克隆等技术获得BcVIL2基因,对其序列进行蛋白分子量(MW)、糖基化位点、二级结构等生物信息学分析;利用荧光定量PCR等技术分析其应答春化表达模式。【结果】BcVIL2基因ORF(Open reading fragment)全长为1296 bp,共编码431个氨基酸,属于亲水性蛋白,预测含有1个潜在的糖基化位点和59个潜在的磷酸化位点,蛋白分子量为47.91 kDa,理论等电点(pI)为8.27;其二级结构主要由α⁃螺旋与无规则卷曲组成;亚细胞定位预测其可能在细胞核中发挥作用;同源序列比对分析显示该基因与芸薹属植物VIL2基因同源性较高;亲缘进化树分析也显示该基因与白菜(B.rapa,syn.B.campestris)和芥菜(B.juncea)VIL2基因亲缘关系很近;荧光定量PCR结果表明,盛花期时BcVIL2基因在根中表达量最低,在荚中表达量最高,而叶中表达量显著低于茎和花中表达量;一月苗龄植株春化后BcVIL2基因的表达量显著低于未春化时的表达量。【结论】乌菜VIL2基因响应春化负表达,与春化反应相关基因BraA01g032910.3C、BraA05g026410.3C与BraA10g016160.3C密切相关,可能存在互作关系,本研究结果为深入分析BcVIL2基因的功能提供了良好基础。

白菜核雄性不育两用系AFLP扩增特异片段的克隆及序列分析

利用AFLP技术从白菜核不育两用系"矮脚黄"可育系DNA中得到一个特异扩增片段MF-14.Southern杂交结果证实不育系也存在该片段的同源序列;回收该特异扩增片段并将其克隆到pGEM-TEasy载体进行序列测定.测序结果表明,该片段全长289bp,其碱基组成为A+T=46.02%.与白菜RAPD特异片段S107及其他育性相关基因核苷酸序列比较,同源性均低于50%.表明该片段为一新的序列.