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.

抗根肿病油菜垦油杂741R的选育及利用潜力

针对湖北省多地出现根肿病危害现象,为抵抗根肿病减小其对油菜(Brassica campestris)生产和推广的影响,提高油菜生产效益,中垦锦绣华农武汉科技有限公司联合华中农业大学利用丰产高抗型稳定细胞质雄性不育系N19A为母本,以具有抗根肿病能力的R741为父本配置组合,育成具有抗油菜根肿病(4号小种)的油菜新品系垦油杂741R。该品系为半冬性甘蓝型油菜萝卜细胞质雄性不育三系杂交种,具有苗期生长势强、分枝性较强、成熟期一致性好、抗倒、抗病等特征。芥酸含量0.1%,硫苷含量23.50μmol/g(饼),符合国家双低标准,含油量45.94%,含油量高,适宜在长江中游生态区湖北省、湖南省、江西省等地推广密植,直播种植密度以37.5万~45.0万株/hm~2为宜。

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。

Study on Agrobacterium tumefaciens-mediated Transformation of Brassica campestris L. with Fusion Gene Ycoil-bFGF

[ Objective] The study is to generate pharmaceutical protein via plant transgenic technique. [Methed] Using the cotyledons with petiole as transformation receptor, the fusion gene of rapeseed oil-body gene and bFGF was introduced into the rapeseed （ Brassica campestris L. ） by Agrobacterium tumefaciens-mediated transformation; meanwhile regeneration conditions of rapeseed were also optimized, and the regenerated resistant plantlets were detected by PCR and Southern blot. [ Result] This fusion gene had been integrated into rapeseed genome successfully, and the optimized conditions of transformation and regeneration were as follows： explants pre-culture for 2 d, co-culture for 3 d, bacteria solution OD600 for 0.3 and infection time for 5 min. [ Conclusion] The results laid a solid foundation for extraction, isolation and purification of protein in transgenic plant seeds.

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.

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.

水稻秸秆还田对油菜产量和土壤理化性质的影响

秸秆还田是维持和改善土壤质量、提高作物产量的有效途径。采用田间试验结合网袋培养的方法,分析了油菜(Brassica campestris)全生育期水稻秸秆的腐解状况,以及水稻(Oryza sativa L.)秸秆还田后油菜产量和土壤肥力特征。结果表明,在52 d时水稻秸秆累积腐解率达13.35%,油菜苗期平均腐解速率为0.26%/d;在229 d时累积腐解率为44.56%,油菜成熟期平均腐解速率为0.14%/d,腐解速率随着腐解时间的延长逐渐趋于平缓。与秸秆不还田相比,秸秆还田显著提高了油菜角果数、角粒数及产量,其中油菜产量提高11.11%;土壤全钾、硝态氮和有效磷含量分别显著增加12.31%、24.55%和25.75%,表明秸秆还田有助于提高土壤中全钾、硝态氮和有效磷含量,保证土壤肥力。相关性分析表明,土壤全氮、全磷、全钾、有机质、硝态氮、有效磷都与油菜产量呈显著正相关(P<0.05)。

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.

Analysis on Cloning and Expression of WRKY Gene Segment in Brassica campestris L.

[Objective]The aim was to explore the relative expression trends of WRKY transcription factor gene under ABA treatment in Brassica campestris.[Method]Actin gene and WRKY gene were cloned by using the homology cloning method.The sequences of nucleic acid and amino acid were analyzed using BLAST and DNAMAN software.Relative expression trends of WRKY gene were detected by applying real-time relative quantification PCR（RT-qPCR）under ABA（100 μmol/L）treatment.[Result]A 680 bp WRKY gene segment and a 933 bp β-actin gene segment were acquired in Brassica campestris.The result of RT-qPCR analysis revealed that BcWRKY expression could be induced by ABA and that the relative expression of WRKY gene reached the peak at 1 h with ABA treatment.[Conclusion]Actin and WRKY gene in B.campestris were cloned successfully,which was proved to play an important role in ABA signal pathway.

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.

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.

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 Genie 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.

Effects of Different Simple Cultivation Facilities on Yield and Quality of Over-summer Brassica campestris L. in South China

The effects of different simple cultivation facilities on yield and quality of Brassica campestris L. were compared to explore the optimal facility type for pro- duction of B. campestris, thereby providing a technical reference for efficient cultiva- tion of B. campestris during hot season. A total of four treatments were designed, open field cultivation （CK）, ordinary anti-insect net shed cultivation, ordinary plastic film greenhouse cultivation and Japanese translucent anti-insect net shed cultivation. The results showed that compared with those in open field cultivation, the plant height, leaf area, yield, Vc content, soluble sugar content and soluble protein con- tent of B. campestris in Japanese translucent anti-insect net shed cultivation were increased by 22%, 63%, 49%, 14%, 13% and 10%, respectively, and Japanese translucent anti-insect net shed was the optimal facility type for cultivation of B. campestris in hot season.

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.

A Genetic Linkage Map of Brassica rapa Based on AFLP Markers

A F2 mapping population was developed by crossing a Chinese cabbage-pe-tsai variety CC156 and an oil type Rapid cycling RC144 which were different from each other in morphology, maturity, self-compatibility, plant height, etc. Using 244 AFLP markers a map was constructed containing 10 main linkage groups covering a total distance of 857 cM, corresponding to 3.5 cM per marker. Length of linkage groups varied from 43 to 125 cM and the number of AFLP markers linkage to each group ranged from 7 to 41.

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.

Interspecific Hybridization of Brassica campestris × B.oleracea Through Ovary Culture

Using three varieties of Brassica campestris, Hauarad (708), Maoshan-3 (714) and Youbai (715),as the maternal plants and one variety of Brassica oleracea Jingfeng-1 (6012) as paternalplants, crosses were made to produce interspecific hybrids through ovary culture techniques.The ovaries from the cross between B. campestrisB.oleracea (7086012 and 7146012) werecultured and ovary culture was more effective in terms of obtained seeds when ovaries werecultured in vitro at 9 d after pollination (DAP). While for the cross of 7156012, it wasbetter when ovaries in vitro cultured at 12 DAP. Among three cross combinations, the cross of7146012 showed the best response and 43 seeds per ovary were obtained. Among the mediastudied, the ovaries from the cross of 7086012 cultured on MS media supplemented with 3.0 mgL-1 BA0.1 mg L-1 NAA showed better response, and its rate of seeds per ovary reached 44.0%.While the ovaries from the other two crosses (7146012 and 7156012) showed the best responsewhen cultured on B5 media supplemented with 3.0 mg L-1 BA + 0.2 mg L-1 NAA, and the rates of seedsper ovary reached 72.0 and 60.0%, respectively. All seeds obtained from the three crosscombinations were cultured on the MS media supplemented with 1.0 mg L-1 BA + 0.05 mg L-1 NAA,and the seeds from the cross of 7156012 showed the best germination response and thepercentage of germinations reached 66.7%. The regenerated plantlets were obtained from theseseedlings after cultured on the MS media supplemented with 0.05 mg L-1 NAA. Cytological studyshowed that these regenerated plants were all true hybrids of B.campestrisB.oleracea.

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.

新型肥料在稻-油轮作制中的施用效果

为探索轮作周期中不同肥料高效施用技术,通过田间定位试验,设置了不施肥(T1)、常规化肥(T2)、缓控释肥(T3)、稳定性肥料(T4)、BB肥(T5)和在T2的基础上增施生物有机肥(T6)6个处理,研究了不同肥料施用对稻(Oryza sativa L.)-油(Brassica campestris L.)轮作制度下作物产量及养分利用效率的影响。结果表明,与施用常规化肥处理相比,施用缓控释肥和在常规化肥基础上增施有机肥均可提高水稻和油菜及周年产量,大部分轮作年份达显著水平(P<0.05);不同肥料处理均能提高稻-油轮作水稻季、油菜季及周年作物氮、磷、钾的吸收量和农学利用效率,其中以T6处理效果最好,氮、磷、钾吸收量和农学利用效率均与施用常规化肥处理差异达显著水平(P<0.05),T3处理效果次之,除油菜季磷吸收量外,其他均与施用常规化肥处理差异达显著水平(P<0.05)。综合来看,湖北省稻-油轮作制中施用缓控释肥和增施生物有机肥效果较好,施用BB肥和稳定性肥料的效果略差。