人工合成甘蓝型油菜特长角变异系的选育

用中国西藏东部采集的1个白菜型油菜地方品种与云南甘蓝类蔬菜白花芥蓝远缘杂交,人工合成了甘蓝型油菜新材料,从杂种后代中通过系统选育,获得了1个特长角变异系,其主花序中部角果平均长度20cm左右,果身长16～18cm,果喙长3cm左右.从中获得的极端最长角果达31.5cm,果身长26.1cm,果喙长5.4cm.这是迄今芸苔属植物中很少见到的长角果油菜材料.该材料的平均角果长度大约为普通甘蓝型油菜的3倍左右,遗传已基本稳定,定名为川农特长角.本文报导其选育经过和主要特征特性,并对其育种和研究利用价值作了简要分析.     

湖北省荆门荆山余脉种子植物区系研究

对湖北省荆门荆山余脉种子植物区系进行了统计分析。结果表明:(1)物种多样性丰富,共有野生种子植物128科、576属、1226种;(2)优势科、属明显;(3)分布区类型多样,地理成分复杂;(4)温带性质明显,热带成分丰富,与热带植物区系有较高的亲缘关系,并表现出一定的过渡性;(5)区划上属于华东植物地区。     

青花菜细胞质雄性不育系线粒体DNA的提取与RAPD分析

利用分子生物学方法提取青花菜不育系及其保持系线粒体的DNA,经琼脂糖凝胶电泳表明所提取的线粒体DNA纯度较高。通过40个引物进行随机引物多态性扩增,结果表明,由引物S66扩增出一条1 000 bp左右的特异条带BMS1000,该条带是不育系m tDNA所特有的,初步推断该特异条带可能与不育性相关。     

辣椒离体培养及再生体系的研究

选用9个辣椒(Capsicum annuumL.)品种(系),研究了不同激素组合、基因型、外植体类型、苗龄和Ag-NO3等因素对外植体不定芽分化和伸长的影响.结果表明,在6-BA/IAA为10∶1配比下,有利于辣椒外植体的分化再生,而6-BA/IAA为3∶1配比下适合于再生芽的伸长;不同品种辣椒的再生能力差别较大,分化率在13.3%~90.0%之间;辣椒子叶再生能力比下胚轴强,是较好的外植体材料;12~16 d苗龄的外植体分化频率较高;添加4mg?L-1AgNO3可使芽分化率平均提高16.9%.通过比较,筛选出了适合于辣椒芽分化的培养基为MB5(MS无机盐 B5有机成分) 5 mg?L-16-BA 0.5 mg?L-1IAA 4 mg?L-1AgNO3,芽伸长培养基为MB5 3 mg?L-16-BA 1 mg?L-1IAA 2 mg?L-1GA3 4 mg?L-1AgNO3,生根培养基为1/2 MS 0.2 mg?L-1IAA 0.1 mg?L-1NAA.     

YS型小麦温敏不育系育性转换基因的cDNA-AFLP分析

对YS型小麦温敏雄性不育系A3017不育与可育条件下不同发育时期的幼穗提取mRNA,进行cDNA- AFLP表达差异分析,64对引物组合获得差异片段1160个。统计分析表明,单核期表达的差异片段数最多,认为这一时期是育性转换的关键时期。对其中12个在不育或可育条件下表达的差异片段进行同收、克隆、测序,经 BLAST序列比对分析表明,可育条件下的4个EST与物质转运和能量代谢过程的铁转运蛋白1和ATP合酶α亚基,基因表达调控的反转录转座子跳跃多聚蛋白和转座子相似序列同源。不育条件下有5个EST与细胞内信号传导的ZCCT转录因子和光敏色素蛋白,基因表达调控的转座酶、染色体浓缩因子和亮氨酸富集蛋白的编码基因同源。结果表明YS型小麦温敏雄性不育系A3017的育性转换与细胞内基因表达调控、物质和能量代谢及信号传导过程有关。     

抗人胎盘酸性铁蛋白单链抗体的构建、表达与鉴定

目的:制备人胎盘酸性铁蛋白(PAF),构建、表达并鉴定抗人PAF单链可变区抗体片段(scFv)。方法:设计以SfiⅠ、NotⅠ为酶切位点、以(Gly4Ser)3为linker的2对引物,从抗人PAF单克隆抗体可变区基因的克隆载体中扩增VH和VL基因,用重叠延伸PCR在VH和VL基因间引入连接短肽,构建VH-linker-VL的scFv基因。经SfiⅠ、NotⅠ酶切后克隆到原核分泌型表达载体pUC19/119上,转化大肠杆菌TG1和筛选后测序验证,IPTG诱导表达,SDS-PAGE鉴定其相对分子质量(Mr)。以人PAF为抗原,scFv为一抗,抗His单克隆抗体为二抗,间接ELISA鉴定其抗体活性。结果:重叠延伸PCR扩增产物经凝胶电泳可见约700bp的条带,DNA序列分析证明2株抗体具有完整的scFv序列,并含有c-myc和His。IPTG诱导阳性菌表达产物经SDS-PAGE鉴定有Mr约27000的显示条带,符合scFv与表达标签融合蛋白的理论值;间接ELISA证明表达产物可与PAF结合。结论:成功构建并表达了抗人PAF单链抗体,为进一步建立检测PAF的间接ELISA奠定了基础。     

Manipulation by culture mixing and elicitation of paclitaxel and baccatin III production in Taxus baccata suspension cultures

Summary Experiments were carried out with Taxus baccata cell lines showing different paclitaxel-producing capacities (between 1.74 and 19.91 mgl^−1) when growing in a selected product-formation medium that specifically stimulated the production of taxane to the detriment of cell growth. Through mixing low-, medial- and high-producing lines, it could be observed that paclitaxel productivity in the resulting mixed lines was clearly higher than the mean productivity of the individual lines before mixing. This suggests that culture components generated by high-producing individual lines within the population might induce paclitaxel production. Although the accumulation of paclitaxel and baccatin III was higher when 100 μM methyl jasmonate was added to the subcultures of the mixed lines, the results indicate that exogenously applied methyl jasmonate was not the first factor to stimulate taxane production. The possible effects of methyl jasmonate elicitation and paclitaxel accumulation on cell viability are also considered.     

Establishment and characterization of an epithelial cell line from thymus of Catla catla (Hamilton, 1822)

A cell line, CTE, derived from catla (Catla catla) thymus has been established by explant method and subcultured for more than 70 passages over a period of 400 days. The cell line has been maintained in L-15 (Leibovitz) medium supplemented with 10% fetal bovine serum. CTE cell line consists of homogeneous population of epithelial-like cells and grows optimally at 28 °C. Karyotype analysis revealed that the modal chromosome number of CTE cells was 50. Partial amplification, sequencing and alignment of fragments of two mitochondrial genes 16S rRNA and COI confirmed that CTE cell line originated from catla. Significant green fluorescent signals were observed when the cell line was transfected with phrGFP II-N mammalian expression vector, indicating its potential utility for transgenic and genetic manipulation studies. The CTE cells showed strong positivity for cytokeratin, indicating that cell line was epithelial in nature. The flow cytometric analysis of cell line revealed a higher number of cells in S-phase at 48 h, suggesting a high growth rate. The extracellular products of Vibrio cholerae MTCC 3904 were toxic to the CTE cells. This cell line was not susceptible to fish betanodavirus, the causative agent of viral nervous necrosis in a large variety of marine fish.     

Strain variability in fatty acid composition of Chattonella marina (Raphidophyceae) and its relation to differing ichthyotoxicity toward rainbow trout gill cells

Lipid profiles of three strains (Mexico, Australia, Japan) of Chattonella marina (Subrahmanyan) Hara et Chihara were studied under defined growth (phosphate, light, and growth phase) and harvest (intact and ruptured cells) conditions. Triacylglycerol levels were always <2%, sterols <7%, free fatty acids varied between 2 and 33%, and polar lipids were the most abundant lipid class (>51% of total lipids). The major fatty acids in C. marina were palmitic (16:0), eicosapentaenoic (EPA, 20:5ω3), octadecatetraenoic (18:4ω3), myristic (14:0), and palmitoleic (16:1ω7c) acids. Higher levels of EPA were found in ruptured cells (21.4–29.4%) compared to intact cells (8.5–25.3%). In general, Japanese N‐118 C. marina was the highest producer of EPA (14.3–29.4%), and Mexican CMCV‐1 the lowest producer (7.9–27.1%). Algal cultures, free fatty acids from C. marina, and the two aldehydes 2E,4E‐decadienal and 2E,4E‐heptadienal (suspected fatty acid‐derived products) were tested against the rainbow trout fish gill cell line RTgill‐W1. The configuration of fatty acids plays an important role in ichthyotoxicity. Free fatty acid fractions, obtained by base saponification of total lipids from C. marina showed a potent toxicity toward gill cells (median lethal concentration, LC₅₀ (at 1 h) of 0.44 μg · mL^?1 in light conditions, with a complete loss of viability at >3.2 μg · mL^?1). Live cultures of Mexican C. marina were less toxic than Japanese and Australian strains. This difference could be related to differing EPA content, superoxide anion production, and cell fragility. The aldehydes 2E,4E‐decadienal and 2E,4E‐heptadienal also showed high impact on gill cell viability, with LC₅₀ (at 1 h) of 0.34 and 0.36 μg · mL^?1, respectively. Superoxide anion production was highest in Australian strain CMPL01, followed by Japanese N‐118 and Mexican CMCV‐1 strains. Ruptured cells showed higher production of superoxide anion compared to intact cells (e.g., 19 vs. 9.5 pmol · cell^?1 · hr^?1 for CMPL01, respectively). Our results indicate that C. marina is more ichthyotoxic after cell disruption and when switching from dark to light conditions, possibly associated with a higher production of superoxide anion and EPA, which may be quickly oxidized to produce more toxic derivates, such as aldehydes.