大白菜(Brassica pekinensis L.)对镉富集基因型差异的研究

研究了15种大白菜基因型对镉(Cd)富集的基因型差异,以期筛选出低积累Cd的白菜基因型.盆栽试验结果表明,不同白菜基因型的Cd含量、富集系数(BFs)和转运系数(TFs)差异显著(P<0.05).丰源新3号、现代夏秋和春秋六轮的Cd积累量均小于0.50mg/kg,其富集系数和转运系数都小于1.0,具有一定低积累Cd的潜力.大田试验的结果表明丰源新3号的Cd含量小于0.50mg/kg,富集系数和转运系数均小于1.0,表明其为排斥Cd的白菜基因型.现代夏秋和春秋六轮的富集系数和转运系数均小于1.0,但其地上部Cd含量均高于0.50mg/kg,不属于Cd低积累白菜.值得注意的.北京小杂56在盆栽和大田中的Cd含量、富集系数和转运系数均大于1.0,可用于植物修复污染土壤.

Pathogen_resistant Transgenic Plant of Brassica pekinensis by Transfering Antibacterial Peptide Gene and Its Genetic Stability

The soft rot infected by pathogenic bacterium Erwinia aroideae Holland is one of the three serious diseases of Chinese cabbage ( Brassica pekinensis Rupr.). By constructing vector system of high frequency transformation mediated by Agrobacterium tunefaciens EHA105, anti-bacterial peptide gene with strong bactericidal action to pathogenic bacteria was introduced into Chinese cabbage AB-81 self-bred line and the transgenic plants were obtained. PCR and Southern blotting detection showed that target gene was integrated into plant genome of Chinese cabbage. The tests of bacteriostasis action of the extract from transgenic plants in vitro, and the assay of disease-resistant of transgenic plantlets in the tube and the pot by perfusing inoculation with pathogenic bacteria showed obvious resistance to soft rot. This resistance can be a stable heredity by genetic analysis of generations of transgenic plants self-bred, separation ratio of its R, was 3:1. The resistance to Km and disease of soft rot was still kept in the R-5. These results indicated the possibility of breeding new varieties of anti-soft rot Chinese cabbage by transgenic plants as parents.

土壤质地对高山娃娃菜(Brassica pekinensis)产量和品质的效应分析

选取云南具有代表性的土壤类型,在昆明设置不同土壤质地对高山娃娃菜(Brassica pekinensis)品质效应的盆栽试验。结果表明,在相同的施肥管理条件下,不同质地的土壤上生长的娃娃菜产量差异显著;在容重较轻、通透性较好的土壤下种植的高山娃娃菜株高、叶片数、结球的紧实度等生长发育情况较好,具有优良的商品外观品质和产量表现;高山娃娃菜中蛋白质含量与土壤中细砂粒和粘粒含量显著负相关,与土壤中粉粒、粗粘粒极显著相关;碳水化合物含量与砂粒显著和极显著相关,与粘粒极显著负相关;膳食纤维与粘粒含量极显著负相关;收获食用部分的NO3-含量与砂粒极显著负相关,与粉粒和粘粒极显著相关;维生素C含量与土壤颗粒没有相关性。从蔬菜产量和安全品质综合来看,砂壤为高山娃娃菜较为适宜的土壤质地类型,其次为容重较轻、通透性较好、有机质含量丰富的中性偏碱的壤土。

Effect of Foliar Application of Amino Acid on the Quality and Enzyme Activity of Flowering Chinese Cabbage(Brassica parachinensis Bailey)

A pot experiment was conducted to study the influences of foliar application of glycine,alanine,lysine,and glutamic acid in 200 mg/kg or 500 mg/kg upon the quality and enzyme activity of flowering Chinese cabbage（Brassica parachinensis Bailey）.The results showed that all the application of these four amino acids could increase the yield of flowering Chinese cabbage,significantly raise the content of soluble sugar,and reduce the accumulation of nitrate.The applications of three other amino acids except alanine can increase the content of soluble proteins and decrease the accumulation of oxalic acid.However,the application of amino acid has insignificant influences on the SPAD number of chlorophyll,and causes the decrease of Vitamin C content.Meanwhile,the application of amino acid can improve the activity of nitrate reductase（NR） and glutamate dehydrogenase（GDH） as well.It shows that the application of amino acid is beneficial to improve ammonia metabolism,reduce the accumulation of nitrate and oxalic acid,increase the content of soluble sugar and soluble proteins,and improve the quality of flowering Chinese cabbage.

Effects of field application of phosphate fertilizers on the availability and uptake of lead, zinc and cadmium by cabbage (Brassica chinensis L.) in a mining tailing contaminated soil

A field trial was conducted to evaluate the reduction of bioavailability of heavy metals including lead (Pb), zinc (Zn) and cadmium (Cd) in a soil contaminated by mining tailings in Shaoxing, Zhejiang, China. Three commercial phosphate (P) fertilizers including phosphate rock (PR), calcium magnesium phosphate (CMP), and single superphosphate (SSP) were applied to the plot at three P application rates, 50, 300, and 500 g/m2 with 9 treatments and control (CK). Plants, water soluble and exchangeable (WE) extra...

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 Nitrogen Fertilizer with Nitrification Inhibitor DMPP (3,4-Dimethylpyrazole phosphate) on Nitrate Accumulation and Quality of Cabbage(Brassica campastris L.ssp. pekinesis)

To assess the effects of N fertilizer ammonium sulphate nitrate [(NH4)2SO4 plus NH4NO3;ASN] with the new nitrification inhibitor (NI) 3, 4-dimethylpyrazole phosphate (DMPP)(ASN+DMPP) on yield, nitrate accumulation, and quality of cabbage (Brassica campastrisL. ssp. pekinesis), two field trials were carried out under various soil-climaticconditions in Jinhua City and Xinchang County, Zhejiang Province of China in 2002.Results showed that DMPP could increase the mean yield by+2.0tha-1 in Jinhua, +5.5tha-1 inXinchang, decrease NO3--N content by -9.4% in Jinhua, -7.3% in Xinchang and improvenutritional quality by increasing vitamin C (VC), soluble sugar, K, Fe, Zn contentssignificantly.

大白菜(Brassica pekinensis)与甘蓝(B.oleracea)的种间试管受精

通过大白菜胚珠与甘蓝花粉的试管授粉和受精得到了杂种种子。杂交一代的植株形态和花粉发育情况及根尖细胞染色体数均表明为种间杂种。杂种植株具有父母本的特征,某些性状介于双亲之间;花粉粒发育不正常;根尖细胞染色体数为2n=19。本研究为其他十字花科植物的远缘杂交中克服不亲和性提供了依据。

A Co-Dominant Marker BoE332 Applied to Marker-Assisted Selection of Homozygous Male-Sterile Plants in Cabbage (Brassica oleracea var.capitata L.)

The dominant genic male sterility （DGMS） gene CDMs399-3 derived from a spontaneous mutation in the line 79-399-3 of spring cabbage （Brassica oleracea var. capitata L.）, has been successfully applied in hybrid seed production of several cabbage cultivars in China. During the development of dominant male sterility lines in cabbage, the conventional identification of homozygous male-sterile plants （CDMs399-3/CDMs399-3） is a laborious and time-consuming process. For marker-assisted selection （MAS） of the gene CDMs399-3 transferred into key spring cabbage line 397, expressed sequence tag-simple sequence repeats （EST-SSR） and SSR technology were used to identify markers that were linked to CDMs399-3 based on method of bulked segregant analysis （BSA）. By screening a set of 978 EST-SSRs and 395 SSRs, a marker BoE332 linked to the CDMs399-3 at a distance of 3.6 cM in the genetic background of cabbage line 397 were identified. 7 homozygons male-sterile plants in population P1170 with 20 plants were obtained finally via MAS of BoE332. Thus, BoE332 will greatly facilitate the transferring of the gene CDMs399-3 into the key spring cabbage line 397 and improve the application of DGMS in cabbage hybrid breeding.

Photosynthetic Excitation Pressure Causes Violaxanthin De-epoxidation in Aging Cabbage （Brassica Oleracea L.） Leaves

The purpose of the present studies was analysis of the age induced changes in photochemical efficiency and xanthophyils cycle pigments of the primary cabbage （Brassica oleracea L. cv. Capitata f. alba） leaves. Photochemical efficiency of photosystem Ⅱ （PS Ⅱ） was studied by a pulse amplitude modulated chlorophyll fluorescence apparatus, chlorophyll concentration was analysis spectrophotometrically and xanthophyll cycle pigments were estimated by high-pressure liquid chromatography （HPLC）. Leaf senescence was accompanied with a decrease both in chlorophylls concentration, the photochemical efficiency and rate constant for PS Ⅱ photochemistry whereas non-photochemical parameters increased. Excitation pressure （1-qP） which is a measure of relative lumen acidification increased by 1.2x in aging leaves. The maximum quantum yield of PS Ⅱ showed no significant change. The level of de-epoxidised xanthophylls increased but the concentration of mono- and di-epoxy xanthophylls decreased in aging leaves. A linear relationship between the excitation pressure and the depoxidation state of the xanthophyll cycle pigments and lutein, during the onset of senescence suggests that excitation pressure can be used as a sensor for monitoring the onset of senescence as well for the de-epoxidation state of the xanthophylls responsible for non-photochemical quenching in stressed leaves.

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.

Effect of Avocado （Persea Americana）, Cabbage （Brassica Oleracea） and Ginger （Zingiber Officinale） on Rat Liver and Thyroid Injuries Induced by CCI4 （Carbon Tetrachloride）

Avocado, Cabbage, and Ginger are a part of a regular human diet and have antioxidant, and antitumor effects. The effect of AVOE （avocado）, GE （Ginger） and CE （Cabbage） extracts separately on liver NO （nitric oxide）, MDA （malondialdehyde）, as well as serum AST （aspartate aminotransferase）, ALT （alanine aminotransferase）, total bilirubin, TC （total cholesterol）, T.G （triglyceride）, HDL cholesterol （high-density lipoprotein）, LDL cholesterol （low-density lipoprotein）, TSH （thyroid-stimulating hormone）, T3 （Triiodothyronine）, T4 （Thyroxine） in rats treated and untreated with CC14 （carbon tetrachloride） was studied. The levels of NO, MDA, as well as serum AST, ALT, total bilirubin, TC, T.G, LDL, and TSH, showed an elevation while, HDL, T3 and T4 showed the decline in rats treated with CC14 as compared to control. Treatment of rats with AVOE and GE pre, during, and post CC14 administration improve NO, MDA, as well as serum AST, ALT, total bilirubin, TC, T.G, HDL, LDL, TSH, T3, T4 as compared to CC14. Treatment of rats with CE pre, during, and post CC14 administration did not improve in the thyroid hormones and lipid profile levels as compared to CC14. These findings suggest that avocado and ginger treatment exerts a protective effect on metabolic disorders by decreasing oxidative stress.

Obtaining and Cytological Identification of a Set of Primary Trisomics in Cabbage

Selection of primary trisomics of the cabbage （Brassica oleracea var.capitata L） forms an important basis for gene chromosome mapping and for other genetic studies. The cabbage self-fertilization line - 9601 was used as material, using the root-tip cell chromosome number and pollen mother cell chromosome number identification and karyotype analysis to select the primary trisomics from the progenies of 3x x 2x in the cabbage. Many aneuploid plants with one or two extra chromosomes were obtained and a set of primary trisomics （Tri-1, Tri-2, Tri-3, Tri-4, Tri-5, Tri-6, Tri-7, Tri-8, and Tri-9, in which the Tri-1 and Tri-4 were from 2n＋2 plants and others from 2n＋ 1 plants） was acquired from these plants. Each trisomic exhibited some unique features, such as plant height, plant type, leaf type, size of flower bud, and inflorescence. The triploid crossing by the diploid is a convenient and effective way to select trisomics in the cabbage.

Isolation and Expression Analysis of Two Cold-Inducible Genes Encoding Putative CBF Transcription Factors from Chinese Cabbage （Brassica pekinensis Rupr）

Two homologous genes of the Arabidopsis C-repeat/dehydration-responsive element binding factors （CBF/ DREB1） transcriptional activator were isolated by RT-PCR from Chinese cabbage （Brassica pekinensis Rupr. cv. Qinbai 5） and were designated as BcCBF1 and BcCBF2. Each encodes a putative CBF/DREB1 protein with an AP2 （Apetal2） DNA-bindlng domain, a putative nuclear localization signal, and a possible acidic activation domain. Deduced amino acid sequences show that BcCBF1 is very similar to the Arabidopsis CBF1, whereas BcCBF2 Is different in that it contains two extra regions of 24 and 20 amino acids in the acidic domain. The mRNA accumulation profiles indicated that the expression of BcCBF1 and BcCBF2 is strongly induced by cold treatment, but does not respond similarly to dehydration or abscisic acid （ABA） treatment. However, the cold-induced accumulation of BcCBF2 mRNA was rapid but short-lived compared with that of BcCBFI. The mRNA levels of both BcCBF1 and BcCBF2 were higher in leaves than in roots when plants were exposed to cold, whereas, salt stress caused higher accumulation of BcCBF2 mRNA in roots than in leaves, suggesting that the organ specificity of the gene expression of the BcCBFs is probably stress dependent. In addition, the accumulation of BcCBF1 and BcCBF2 mRNAs was greatly enhanced by light compared with darkness when seedlings were exposed to cold. It is concluded that the two BcCBF proteins may be involved in the process of plant response to cold stress through an ABA-independent pathway and that there is also a cross-talk between the light signaling conduction pathway and the cold response pathway in B. pekinensis as in Arabidopsis.

白菜黑斑病菌（Alternaria brassicae）菌丝生长和毒素产生条件的研究

对白菜黑斑病菌（Ａｌｔｅｒｎａｒｉａｂｒａｓｓｉｃａｅ）不同菌株的营养生长和产毒条件的研究结果表明，白帮菌系（ＡＢＷ）与青帮菌系（ＡＢＧ）的生物学特性有一定的差异，同一菌系菌株间差异不显著，其中ＡＢＷ菌丝生长的最佳条件是２０～２２℃、ｐＨ４、光照１２ｈ／ｄ、间歇振荡和以蔗糖（浓度为３％）为Ｃ源；而毒素产生的最佳条件是１３～１５℃、ｐＨ６、黑暗、静止和以葡萄糖（浓度为１０％）为Ｃ源。ＡＢＧ菌丝生长的最佳条件是１３～１５℃、ｐＨ４、光照１２ｈ／ｄ、全天振荡和以蔗糖（浓度为３％）为Ｃ源；而毒素产生的最佳条件是１３～１５℃、ｐＨ４、黑暗、静止和以葡萄糖（浓度为５％）为Ｃ源。研究还发现，相同条件下ＡＢＧ的菌丝生长速度均大于ＡＢＷ，而产毒量却较为复杂。青帮品种比白帮品种白菜更易感染白菜黑斑病菌。此外，白菜黑斑病菌生长后的改良Ｆｒｉｅｓ培养基ｐＨ值均有变小的趋势，而且随着菌丝生长量的增加变化幅度逐渐增大。

根癌农杆菌介导白细胞介素-4基因转化甘蓝(Brassica oleracea L.)研究

利用根癌农杆菌介导法，以白细胞介素-4（interleukin-4，IL-4）基因转化中甘11号甘蓝。本研究对可能影响il-4转化率的外植体类型、外植体的预培养时间、农杆菌的侵染时间以及外植体与农杆菌的共培养时间进行了优化。试验结果表明：il-4基因对带1-2mm子叶柄的子叶的转化率显著高于下胚轴；带柄子叶在预培养1d、侵染3min、共培养1d时，转化效果最好，转化率达23．3%;下胚轴在预培养1-3d，侵染3～5min，共培养1-2d时，转化效果较好，转化率最高可达13．3％。经PCR检测及PCR-Southem杂交，初步证明目的基因il-4已经转入甘蓝的再生植株。转il-4基因甘蓝的PCR阳性再生植株已经开花并产生了后代。

芸薹根肿菌(Plasmodiophora brassicae Wor.)DNA提取方法的研究与应用

芸薹根肿菌是专一的活体寄生菌,通过分子技术对其群体遗传结构研究的过程中,根肿菌DNA的提取是基础之一。利用硅藻土法和改良CTAB法从根肿菌孢子中直接提取芸薹根肿菌DNA,使用PCR技术对所得到的DNA用大白菜特异引物TCR01、检测根肿菌特异引物TC01进行检测,对2种根肿菌DNA的提取方法进行比较,采用EST-SSR分子标记对纯单孢菌系4号、10号生理小种及混合菌1,2,3,4,7,10,11,14,16号生理小种进行分析。结果表明:硅藻土法操作步骤少,较改良CTAB法操作简单;且硅藻土法提取获得DNA的PCR结果优于CTAB法,能够隔离寄主DNA。EST-SSR标记能对单孢菌4号、10号生理小种特异扩增,片段大小分别约为200bp,混合菌4号、10号生理小种扩增结果相符。H2的带型与H1中的1条带型一致,而H2、H7、H10、H11、H14和H16主带型基本一致,不同生理小种的根肿菌的变异性和致病性可能是相似的。

水肥耦合对娃娃菜叶球养分吸收和土壤理化性质的影响

为探明河西冷凉灌区秋茬娃娃菜节水节肥、高产高效的科学灌溉施肥制度,以‘金皇后’为试材,研究了膜下滴灌条件下3个灌溉水平:60%θ_(f)(低水,H1)、70%θ_(f)(中水,H2)、80%θ_(f)(高水,H3)和3个施肥水平:N 281-P_(2)O_(5)166-K_(2)O 383 kg·hm^(-2)(低肥,F1)、N 330-P_(2)O_(5)195-K_(2)O 450 kg·hm^(-2)(中肥,F2)、N 380-P_(2)O_(5)224-K_(2)O 518 kg·hm^(-2)(高肥,F3)组合对娃娃菜叶球养分吸收和土壤理化性质的影响。结果表明:水肥用量与娃娃菜田土壤理化性质和叶球氮磷钾吸收量关系密切,H2F2处理可在一定程度上改善娃娃菜田土壤的理化性质,维持土壤酸碱平衡,提高叶球养分含量,增加养分累积量。水肥耦合模式下,H2F2处理较H3F3处理娃娃菜叶球氮、磷、钾吸收量分别提高4.72%、3.26%、3.22%。H2F2处理水平下基于隶属函数与标准差系数权重法取得的水肥耦合效应综合得分最高,且较H3F3处理提高7.13%。综合考虑娃娃菜田土壤理化性质和叶球氮磷钾吸收量,灌水下限控制在70%θf、施肥量N 330-P_(2)O_(5)195-K_(2)O 450 kg·hm^(-2)是适合甘肃河西走廊冷凉灌区露地膜下滴灌秋茬娃娃菜最优的水肥管理模式。

不结球白菜BcCHC1蛋白参与调控TuMV侵染的初步研究

【目的】为增强不结球白菜对芜菁花叶病素养(TuMV)的抗性,研究探讨了不结球白菜中BcCHC1蛋白与TuMV之间的相互作用机制。【方法】首先从不结球白菜中鉴定出重链网格蛋白(CHC)基因家族的成员BcCHCs,随后克隆了1个代表性的CHC1基因,并对其进行了亚细胞定位分析。通过双分子荧光互补实验(BiFC)筛选出与BcCHC1蛋白相互作用的TuMV蛋白,并利用病毒诱导的基因沉默技术(VIGS)来沉默BcCHC1基因,以评估其功能。【结果】(1)成功克隆了不结球白菜的BcCHC1基因,该基因的编码序列长5124碱基对,编码1708个氨基酸。(2)在TuMV感染不结球白菜30 d后,实时荧光定量PCR分析显示,感染TuMV的不结球白菜中BcCHC1基因的表达量显著降低。(3)亚细胞定位研究表明,BcCHC1蛋白主要定位于烟草表皮细胞的细胞膜和细胞核。(4)BiFC实验进一步证实,BcCHC1蛋白能够与TuMV的CI和6K2蛋白发生相互作用,其中与CI的互作主要发生在细胞核内,而与6K2的互作则主要位于细胞膜。(5)通过对BcCHC1基因沉默株系的观察发现,沉默BcCHC1基因会导致植株死亡。【结论】BcCHC1蛋白通过与TuMV的CI和6K2蛋白相互作用,影响网格蛋白依赖的内吞作用途径及病毒复制,从而调控TuMV对不结球白菜的侵染。然而,BcCHC1基因调控TuMV侵染不结球白菜的具体机制仍需进一步研究。