Effect of Combined Use of Boron and Manganese Fertilizers on the Nutritional Quality and Physiological Indices of Brassica campestris L.ssp.chinensis L.var.utilis

[Objective] The purpose of this study is to determine the effects of com-bined use of boron and manganese fertilizers on the nutritional quality and physio-logical indices of Brassica campestris. [Method] In the nutrient solutions for growing B. campestris by hydroponics, boric acid and manganese sulfate were added at 0.5, 2.5, and 7.5 mg/L respectively. Another treatment without boron and manganese was prepared as the control. Quality and physiological indices of B. campestris in the 10 treatments were measured. [Result] Boron and manganese shared obvious in-teraction in improving the quality and physiological indices of B. campestris. To cul-tivate B. campestris with high quality and strong resistance, the optimum concentra-tions of boron and manganese in the nutrient solution should be 2.5 mg/L boric acid and 2.5-7.5 mg/L manganese sulfate. [Conclusion] The findings wil provide refer-ence for studying effects of trace elements on nutrient composition of vegetables.

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.

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.

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.

不结球白菜(Brassi cacampestris ssp.chinensis Makino)种质资源SRAP遗传分化分析

利用SRAP分子标记分析了国内外64份不结球白菜种质资源的DNA遗传多样性和遗传分化。21对引物组合共检测出215个位点,其中112个为多态性位点,多态性比率达52.09%,平均每对引物组合产生10.24个位点和5.33个多态性位点。不结球白菜各类型中普通白菜的Nei’s基因多样性指数(0.1410)和遗传丰富度[(190)88.37%]最高;各生态区域中江淮流域不结球白菜的Nei’s基因多样性指数(0.1451)和遗传丰富度[(185)86.05%]最高;国内的Nei’s基因多样性指数(0.1293)和遗传丰富度[(188)87.44%]分别高于国外。遗传变异估算表明,不结球白菜遗传分化系数58.22%,大部分变异存在于种群间;基因流为0.4031,说明群体间基因流动较少,遗传分化程度较高。以遗传相似系数0.872为截值,可把不结球球白菜分为Ⅵ个类群。

Research Progress of Heat-tolerance of Brassica campestris ssp. Chinensis

Brassica campestris ssp. chinensis, also known as non-heading Chinese cabbage, is an important vegetable widely distributed in southern China. High temperature is the most common adversity factor in vegetable production, because Brassica campestris ssp. chinensis is a thermophilic vegetable, which can＇t well grow at high temperature. In summer and autumn, high temperature stress would prevent Brassica campestris ssp. chinensis from growing and result in poor quality of its seedlings, seriously influencing yield and quality of the vegetable in later period. In this paper, present situation and latest advances in heat resistance research of Brassica campestris ssp. chinensis in recent years were reviewed at home and abroad. The research tendency in Bassica campestris ssp. chinensis was also described, providing reference for breeding of heat-resistant Brassica campestris ssp. chinensis.

Genetic Inheritance and Molecular Marker of Clubroot Resistance Genes in Brassica campestris ssp. chinensis

Objective] This study was conducted to investigate the genetic inheritance of clubroot resistance in Chinese non-heading cabbage （Brassica campestris ssp. chinensis）. [Method] The clubroot resistance gene was introduced from a Brassica campestris ssp. pekinensis cultivar to non-heading Chinese cabbage, and the inheri-tance and molecular markers of clubroot resistance gene in parental lines, F1, F2 and BC1 of non-heading Chinese cabbage were studied through pathogen inoculation at seedling stage and ISSR-PCR. [Result] Clubroot resistance in non-heading Chi-nese cabbage was control ed by a single dominant gene. ISSR molecular markers with Bulk segregant analysis （BSA） found that primer-873 was linked to resistance gene, named CR-873, and the genetic distance between the marker and the resis-tance gene was 9.72 cM. [Conclusion] The results provide references for the molecular marker assisted breeding of non-heading Chinese cabbage.

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.

Measurement of Some Physiological and Biochemical Indexes of the Antisense Fragment of CYP86MF Gene Transgenic Male Sterile Plantlets in Chinese Cabbage-Pak-Choi

In order to study the mechanism of the transgenic male sterility induced by the antisense fragment of gene CYP86MF, some physiological and biochemical indexes were compared between the transgenic plantlets of Chinese cabbage-pak-choi(Brassica campestris L. ssp. chinensis Makino)and their controls. Results showed that there was significant difference between content of the endogenous hormones in leaves and floral buds. GA3 contents in leaves of the transgenic plantlets of B. campestris L. ssp. chinensis Makino cv. Shanghai-qing and cv. Youqing were reduced by 31. 72 and 46. 88% respectively as compared with those of their controls, and GA3 contents in floral buds of them were reduced by 62. 92 and 80. 57 % respectively. In addition, ZT contents in leaves of cv. Shanghai-qing and cv. Youqing were reduced by 11. 81 and 181. 20% respectively as compared with those of their controls and those in floral buds were reduced by 105. 94 and 128.75 % respectively. But, ABA content in transgenic plantlets was significantly higher than that in non-transgenic plantlets. However, differences among respiratory rate and cyanide-resistant respiratory rate, the photosynthesic rate and the content of total flavones in floral buds were not significant. In this study, the results indicated that the gene CYP86MF might be one of the key functional genes to control fertility of Chinese cabbage.

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

A genetic linkage map comprised of 131 loci was constructed with an F 2 population derived from an inter subspecific cross between Brassica campestris L. ssp. chinensis cv. 'aijiaohuang' and ssp. rapifera cv. 'qisihai'. 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.

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.

Chinese Cabbage-pak-choi Transcriptome Map Construction with cDNA-AFLP Techniques

Chinese cabbage-pak-choi （Brassica campestris L. ssp. chinensis Makino） transcriptome map with cDNA-AFLP techniques was constructed. The inbred line Aijiaohuang 97-3-2, the inbred line Baimanjing 001-24 of turnip [B. campestris ssp. rapifera （Matzg.） Sinsk] and 183 F6 （recombinant inbred population） plants were used as experimental materials, cDNAs were synthesized from total RNA extracted from young leaves at rosette stage. 256 pairs of cDNA-AFLP primers were used to detect the polymorphisms between parents Aijiaohuang 97-3-2 and Baimanjing 001-24. 56 pairs of cDNA-AFLP primers with high polymorphisms were screened from 256 pairs of primer by DNA-AFLP techniques. The genetic diversity of parents and 183 F6 progenies was detected by 56 pairs of cDNA-AFLP primers. The segregation and distribution of cDNA-AFLPs molecular marker were analyzed to construct transcriptome map amongst parents and F6 plants. A total of 164 cDNA-AFLPs marker loci were mapped into 13 linkage groups which covered 1401.2 cM with an average distance of 9.7 cM. It was the first transcriptome map of Chinese cabbage using cDNA-AFLP technique.

Preliminary Study on Tissue Culture Technique of Brassica campestris L. ssp. chinensis (L.) Makino var. utilis

In order to establish a rapid propagation system for Brassica campestris L. ssp. chinensis var. utilis, the anther and cotyledon-cotyledonary petioles were used as explants to conduct tissue culture research. The results showed that not bloomed flower buds with higher styles than petals were appropriate for anther cul- ture. Moreover, most microspores were at the mid-late uninucleate stage at that time. The pollen germination rates of Brassica campestris L. ssp. chinensis var. u- tills were low and the germination rates in autumn and winter were higher than that in summer. The induction medium of anther callus of Brassica campestris L. ssp. chinensis var. utilis was MS＋1.0 mg/L KT＋1.0 mg/L 2, 4-D＋3% sugar＋6 g/L agar＋ 8% coconut milk and the induction media of adventitious bud were MS＋2.0 mg/L 6-BA＋ 0.5 mg/L NAA＋I.0 g/L activated carbon＋2% sugar＋6 g/L agar or MS＋2.0 mg/L ZT＋ 0.5 mg/L IAA＋0.5 g/L AgNO3＋1.0 g/L activated carbon＋2% sugar＋6 g/L agar. The induction percentage of adventitious bud by anther culture was 36.7%. Browning phenomenon appeared during the culture of adventitious bud and regeneration plant could not be formed. The plant regeneration rate reached 80% when cotyledon- cotyledonary petioles were used as explant.

不结球白菜GST基因家族的全基因组鉴定及花青素相关基因Bc GSTF6的功能分析

【背景】花青素是紫色不结球白菜中重要的营养成分之一,谷胱甘肽转移酶(glutathione S-transferase,GST)在花青素转运中发挥着重要功能。【目的】鉴定不结球白菜中编码GST的基因,并通过转录组和分子生物学手段揭示BcGSTF6在不结球白菜花青素转运中的作用,解析不结球白菜花青素积累的分子机制。【方法】通过生物信息学方法,鉴定GST基因家族成员并分析其在染色体上的位置和基序;在紫色和绿色的不结球白菜中进行转录组分析,筛选不结球白菜中参与花青素转运的基因,并通过表达模式分析、亚细胞定位、拟南芥突变体tt19的异源回补试验进一步验证BcGSTF6和BcTT19的功能;通过将BcGSTF6和BcTT19与其他物种中参与花青素转运基因的氨基酸序列进行比较,分析BcGSTF6存在的差异;并通过BcGSTF6蛋白与矢车菊素(Cya)的体外结合和基因沉默试验验证BcGSTF6是否为参与不结球白菜花青素转运和积累的重要基因。【结果】从不结球白菜中鉴定得到83个GST基因家族成员,分为8个亚家族,分布在10条染色体上。转录组分析结果表明BcGSTF6和BcTT19可能是参与不结球白菜花青素转运的基因,进一步利用qRT-PCR发现BcGSTF6与BcTT19都在不结球白菜花青素积累阶段高度表达;亚细胞定位结果表明BcGSTF6和BcTT19都定位于内质网和液泡膜中。氨基酸序列多重比较发现BcTT19与其他物种中参与花青素转运的蛋白存在高度同源性,而BcGSTF6则在保守结构域中存在较大差异。在拟南芥突变体tt19中对BcGSTF6和BcTT19进行过表达均可恢复拟南芥幼苗tt19积累花青素的表型,但不能恢复其种皮棕色的表型,表明BcGSTF6和BcTT19都参与了花青素的转运但不能转运原花青素(PAs)。体外结合试验发现BcGSTF6蛋白在室温下可增加Cya的可溶性;对BcGSTF6在不结球白菜中进行基因沉默发现叶片中花青素含量显著下降,同时也导致参与花青素合成与调控相关基因表达量降低。证明BcGSTF6参与了花青素的转运和积累,在不结球白菜叶片着色过程中发挥重要功能。【结论】本研究鉴定了不结球白菜GST基因家族,并进一步解析了BcTT19和BcGSTF6在不结球白菜花青素转运和积累中的功能,阐明了BcGSTs在不结球白菜中花青素转运调控的部分机制。

不结球白菜细胞质雄性不育材料的筛选及其保持系选育

用多种不同基因型的可育不结球白菜(Brassica campestris ssp. chinensis Makino)材料对2个不结球白菜雄性不育株进行授粉,根据F1代群体的育性表现,筛选出细胞质雄性不育株;对16个自交品系的纯度进行调查,各品系筛选出5个或10个典型植株对其株高、开展度、叶长、叶宽、叶柄长、色泽等性状进行测试和比较分析。结果表明,16个品系的纯度均不低于50%;1-5、3-31、2-16、2-61、1-79、华冠4的开展度、株高均较大;聚类结果显示,3-31、1-9、2-61、2-16和华冠4聚在第三类,品系间性状差异较小。1-5、1-79、3-31、2-61、1-26、2-11、2-25和1-98各性状的变异系数均低于30%,一致性好;16个自交品系叶色空间分布较叶柄色集中,说明品系间叶色差异较小;3-31、1-79、1-34-17、2-11、2-16、1-5和华冠4的叶色亮丽。综合分析,1-5、1-79、3-31和2-11色泽好,变异系数小,且纯度高,是选育保持系的优良品系;而华冠4和墩子黄既可以作保持系又可以作父本。3-31和华冠4的植株高大,分别适宜作为保持系和父本,用于选配高产的不结球白菜杂交组合。

Cloning and Characterization of the Microspore Development-Related Gene BcMF2 in Chinese Cabbage Pak-Choi （Brassica campestris L. ssp. chinensis Makino）

: For the sake of providing some important information relevant to the study of the molecular mechanism of genic male sterility in plants, gene differential expression in flower buds at different developmental stages, as well as in rosette leaves, florescence leaves, and scapes was analyzed using cDNA amplified fragment length polymorphism (cDNA-AFLP) in the genic male sterile A and fertile B line of Chinese cabbage pak-choi. Following amplification of 125 pairs of primer combinations, 11 differential fragments were obtained, of which eight were from the B line and the other three were from the A line. Of 11 differential fragments, four were verified by Northern hybridization that were expressed preferentially in fertile flower buds. Results of GenBank BLAST showed that one fragment was with unknown function, whereas the other fragments have strong nucleotide sequence similarities with the polygalacturonase (PG) gene, the pectinesterase (PE) gene, and the polygalacturonase inhibitory protein (PGIP4) gene. Only full-length cDNA from the differential fragment BcMF-A18T16-1 was amplified by rapid amplification of cDNA ends (RACE) and Northern analysis showed that this fragment was expressed only in medium and large-sized flower buds of the B line. The full-length cDNA, designated as BcMF2 (Brassica campestris Male Fertile 2), was 1 485 bp long and was composed of a 1 263-bp open reading frame, which had 83% nucleotide similarity to a PG gene from Arabidopsis encoding polygalacturonase. Analysis of the basic structure of the protein revealed that it had one polygalacturonase active site (RVTCGPGHGLSVGS) at 256th site of amino acids and was classified as being a member of family 28 of the glycosyl hydrolases. The role of the BcMF2 gene on microspore development is discussed in the present paper.

镉对小白菜光合作用特性影响的研究

研究表明,微量镉(Cd)(0.01、0.1 mg/L)促进小白菜(Brassica campestris ssp.chinensis)生长,对苏州青的促进作用更明显;高浓度Cd显著抑制地上部生长,对根系生长影响相对较小.Cd降低小白菜叶绿素含量,特别是苏州青降低幅度较大.低浓度Cd对小白菜净光合速率Pn影响不大,高浓度Cd可能是由于非气孔限制而引起Pn降低.Cd降低气孔导度,但高浓度Cd增加胞间CO2浓度.叶绿素荧光测定表明,Cd对小白菜叶片光化学效率Fv/Fm影响不大,而高浓度Cd明显降低光合电子传递量子效率PSⅡ、光化学淬灭系数qP和电子传递速率ETR.Cd增加沪青一号非光化学淬灭系数qNP,而苏州青在高Cd浓度下qNP下降.

白菜和芜菁Ogura型雄性不育系与保持系的获得及其细胞学观察

通过甘蓝型油菜Ogura细胞质雄性不育材料与白菜、芜菁的种间杂交和连续回交,获得了不育性稳定的白菜、芜菁Ogura细胞质雄性不育系‘新选Ogu28-4’、‘矮脚黄Ogu33-14’和‘耐病Ogu18-4’及其相应的保持系。细胞形态学研究表明:虽然这3个不育系核背景不同,但花药败育时期基本相同,均具有单核败育型的特征;小孢子到单核期后不再向前继续发育,而是迅速退化以至解体,开花前彻底败育;不育系的花药在减数分裂早期绒毡层细胞中已出现液泡,有肥大趋势,而正常花药绒毡层的发育在减数分裂过程中达到高峰,四分体时出现退化迹象,开花前完全解体。不同的眙Ogura细胞质雄性不育系,虽然核背景不同,但花药败育的时期基本相同,说明Ogura CMS胞质互作关系稳定,受核背景影响小。

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.

正交设计优化菜薹ISSR反应体系研究

菜薹原产中国,是华南地区重要的特产蔬菜之一。目前,分子标记技术已广泛应用于多种作物研究,但在菜薹上的应用研究刚刚起步。ISSR主要是由单一引物且以重复序列为主要引物序列的PCR标记,其反应条件受模板DNA、Taq DNA聚合酶、Mg2+、dNTP和引物等因素的影响。正交试验设计能明确各因素间的互作效应,建立最优化的反应体系。本研究利用正交试验设计,从Taq DNA聚合酶、dNTP、引物、Mg2+4种因素3个水平对菜薹ISSR反应体系进行了优化,确立了适合菜薹的ISSR反应体系并在7个菜薹品种中进行了验证。在20μl反应体系中,含Taq DNA聚合酶1U、dNTP250μmol/L、引物0.25μmol/L、1×PCRbuffer、Mg2+2.5mmol/L、模板DNA30ng。通过梯度PCR测验,确定了适宜的退火温度。这一体系的建立为今后利用ISSR技术进行菜薹种质资源分类、遗传图谱构建和基因定位奠定了技术基础。