小麦穗突变体Sda1与其野生型叶片总蛋白双向电泳体系的建立

为探索适合小麦穗突变体Sda1叶片蛋白的双向电泳体系,寻找Sda1与其野生型叶片的差异蛋白,以Sda1与其野生型的抽穗期叶片为材料,从蛋白质提取、双向电泳条件等方面对适合Sda1及其野生型叶片蛋白的双向电泳体系进行探索。分析试验得到的双向电泳图谱,发现利用TCA/丙酮提取叶片总蛋白,用pH 4~7的17cm线性胶条,采用13%浓度的分离胶,上样量为900μg,利用改良的等电聚焦程序,能够得到背景清晰、分辨率高的电泳图谱;电泳图谱在低分子区蛋白点分布均匀,并且重复性好。利用PDQuest 8.0.1软件分析图谱,得到27个差异点蛋白,相比于野生型植株,Sda1突变体表现为蛋白表达量下调与缺失,发现6个缺失蛋白,21个表达下调蛋白。     

Contribution of common wheat protein fractions to dough properties and quality of northern-style Chinese steamed bread

Thirty-three cultivars and advanced lines originated from China, Mexico, and Australia were sown in four environments in Chinese spring wheat regions to investigate the association between gluten protein fractions determined by reversed-phase high-performance liquid chromatography (RP-HPLC), and dough properties and northern-style Chinese steamed bread (CSB) quality. The genotypes were divided into two groups of 10 and 23 entries with and without the 1B/1R translocation, respectively. 1B/1R translocation lines had significantly high amounts of ω  -gliadins, and low levels of glutenin and low molecular weight glutenin subunits (LMW-GS), but no significant difference in dough properties and CSB quality from non-translocation lines. The association between protein fractions and dough properties, and CSB quality largely depended upon the presence of 1B/1R translocation. Gliadin contributed more in quantity to flour protein content (FPC) than glutenin, while glutenin and its fractions contributed more to dough strength and CSB quality. Among non-translocation lines, moderate to high correlation coefficients between quantified glutenin and its fractions, and farinograph development time (DT, r=0.85$r=0.85$–0.92) and stability (ST, r=0.81$r=0.81$–0.93), extensograph maximum resistance (R_max, r=0.90$r=0.90$–0.93), CSB stress relaxation (SR, r=0.55$r=0.55$–0.61) and CSB score (r=0.56$r=0.56$–0.62), were observed. Gliadin:glutenin ratios showed significant and negative associations with dough properties and CSB quality. Correlation coefficients between gliadin:glutenin, gliadin:HMW-GS, gliadin:LMW-GS ratios, and CSB score were −0.79, −0.73, and −0.79 among non-translocation lines, respectively. HMW-GS and LMW-GS, x-type HMW-GS and y-type HMW-GS contributed similarly to dough properties and CSB quality for non-translocation lines. Weak correlations between protein fractions and dough properties, and CSB quality were observed among translocation lines. This information should be useful for improvement of dough properties and CSB quality.     

水稻抗病蛋白质XA21的免疫纯化及双向电泳分离研究

在优化水稻叶片蛋白质提取方法的基础上,利用免疫沉淀对水稻中的XA21蛋白质进行了富集纯化,继而通过TCA-丙酮法脱盐,用尿素、硫脲溶解后进行双向电泳分离和Western检测,实现了对高分子量、低丰度表达的、可能的膜蛋白XA21的分离检测,为其翻译后修饰方式研究奠定了基础.     

橡胶树死皮病黄色体蛋白质组差异分析与初步鉴定

应用双向凝胶电泳技术（two—dimensional gel electrophshiya／oresis,2-DE）,比较橡胶树死皮株与健康株胶乳黄色体蛋白质组表达的差异。采用TCA／丙酮沉淀法提取橡胶树死皮株与健康株黄色体蛋白质,并采用固相pH梯度（immobilized pH gradient,IPG）双向凝胶电泳分离两种材料蛋白质,凝胶经银染显色后,用PDQuest图像分析软件进行比较分析、识别差异表达的蛋白质。成功获得了橡胶树死皮株与健康株胶乳黄色体的双向凝胶电泳图谱。鉴定出24个蛋白差异点,其中17个上调表达,7个下调表达。并应用质谱技术鉴定了其中部分表达差异的蛋白质点,对渗调蛋白进行了功能分析,其在死皮株中表现下调的情形可能与死皮病的发生有一定的关系。     

兴凯湖翘嘴红鱼 白同工酶的研究

用水平淀粉电泳技术 ,对兴凯湖翘嘴红鱼白的肌、肝、眼、心 4种不同组织的 8种同工酶 (LDH、EST、MDH、IDH、SOD、ME、G - 6-PDH、ADH)进行了研究。分析了各种同工酶的基因表达谱式。其中 ,在所检测到的 2 0个基因座位中只有EST同工酶中的一个座位 (Est - 2 )表现为多态。兴凯湖翘嘴红鱼白的多态率 (P)为 0 0 50 0 ,平均杂合度 (H)为 0 0 139。与其它鲤科硬骨鱼类相比 ,兴凯湖翘嘴红鱼白在蛋白质水平上的遗传变异表现比较贫乏     

簇毛麦与普通小麦杂种后代稳定品系贮藏蛋白变异分析

为了研究簇毛麦基因向小麦中的导入情况,对151个簇毛麦与绵阳11杂种后代稳定品系贮藏蛋白进行了分析。结果表明在杂种后代的高分子谷蛋白亚基中,除具有小麦亲本绵阳11的亚基外,还存在大量的变异,包括两亲本都不具有,如2+12,9等亚基,但在后代中未发现簇毛麦高分子谷蛋白亚基,说明在杂种后代中存在大量的变异与重组亚基。结果还表明后代中有少量的高分子谷蛋白亚基位点未纯合,因此有必要进一步在后代中进行选择。而醇溶蛋白结果表明在其α、β、γ、ω区都有簇毛麦的谱带和新的谱带出现,说明簇毛麦的一些控制纯溶蛋白的位点已转入小麦中,丰富了小麦遗传多样性。利用醇溶蛋白谱带数据聚类分析表明,后代品系与亲本之间存在较大变异。因此,在远缘杂交后代中不仅可以转移外源基因,而且还可能产生一些新的变异,为进一步研究提供了材料。     

DGGE技术优化及其在肠道微生态研究中的应用

变性梯度凝胶电泳(DGGE)是一种非培养微生物研究方法,克服了实验室传统的微生物分离、培养和分类方法的局限性,是研究动物肠道微生态群落的重要手段之一。介绍了DGGE的技术原理和系统优化及其在动物肠道微生态研究中的应用和自身存在的局限性,并对其应用前景进行了展望。     

鹤顶兰胚珠发育过程及受精前后同工酶的变化

 通过光学显微镜观察发现：鹤顶兰[Phaius tankervilliae (Aiton) Bl.]授粉10 d出现指状突起,20 d形成胚珠原基,24 d大孢子母细胞发生,30 d功能大孢子形成,41 d发育为成熟的八核胚囊,而后完成双受精发育成早期球形胚。用聚丙烯酰胺凝胶电泳的方法研究鹤顶兰从指状突起出现到早期球形胚形成的全过程中酯酶（Est）、酸性磷酸酶（Acp）的变化发现：这一过程中酯酶（Est）表达有7个同工酶位点,酸性磷酸酶（Acp）有5个同工酶位点,两者均在八核胚囊时期变化最显著。Est-7和Acp-4在整个发育过程中都有表达;在八核胚囊时期特异表达的位点有Est-4和Acp-3;而Acp-2只在这一时期不表达。Est-1在八核胚囊时期和大孢子母细胞减数分裂I中期表达。授粉后Est和Acp的酶位点变化说明与胚珠发育相关的一系列基因表达有时序性,位点Est-4、Acp-2和Acp-3是否直接参与了八核胚囊发育的控制,有待于进一步的研究。     

蓝变真菌引起的欧洲云杉木质部解剖学特征及纤维素酶活性的变化

选择奥地利境内阿尔卑斯山健康欧洲云杉为对象,研究室内接种蓝变真菌(Ceratocystis polonica)引起的寄主树木韧皮部和木质部解剖学特征的变化,揭示蓝变真菌引起欧洲云杉枯萎的机制.结果表明:接种1周后的4株欧洲云杉的木质部组织内,蓝变区域显著增加,4～6周后蓝变区域不再增加;而在接种无菌琼脂的2株对照欧洲云杉的木质部组织内,没有检测到蓝变区域.采用生物化学分析和组织定位技术,确定接种真菌和无菌琼脂的欧洲云杉木质部区域纤维素酶的分布和活性变化.接种2周后剥皮取样检测,接种真菌的4株欧洲云杉的木质部组织内纤维素酶含量大幅度增加,其等电聚焦电泳显示明显的纤维素酶酶带;而在接种无菌琼脂的欧洲云杉木质部区域,纤维素酶含量分布较少,其等电聚焦电泳显示微弱的纤维素酶酶带.进一步证明蓝变真菌分泌的纤维素酶能利用寄主欧洲云杉木质部的纤维素,蓝变真菌是致死阿尔卑斯山境内欧洲云杉的重要病原菌.     

Evaluation of protein hydrolysis in raw sources by digestive proteases of Senegalese sole (Solea senegalensis,Kaup 1858) using a combination of an in vitro assay and sodium dodecyl sulphate polyacrylamide gel electrophoresis analysis of products

The hydrolysis of protein in different animal and plant sources by the intestinal proteases of Senegalese sole (Solea senegalensis) was studied using a combination of an in vitro digestibility assay and the evaluation of the protein fractionation by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS‐PAGE). In vitro hydrolysis was performed for 90 min in a closed reactor maintained at constant pH and temperature. Samples of the reaction mixture at different time intervals were developed in SDS‐PAGE gels to evaluate the progressive hydrolysis of the different protein fractions in each protein source. A numerical value [coefficient of protein degradation (CPD)], which integrates the information obtained after image analysis of the gels, is proposed for comparison among proteins, according to the intensity of the hydrolysis produced by sole proteases. Additionally, the total free amino acid released from proteins was measured during the in vitro assay. Casein, squid meal and soybean concentrate (SBC) proteins showed very similar degradation patterns, with a quick and almost complete proteolysis within the first minutes of the enzymatic reaction. Fish and krill meals were hydrolysed more progressively. On the contrary, pea meal (PM) and corn gluten meal (CGM) showed scarce changes in their protein profile after 90 min of reaction. For animal protein sources, the final CPD values ranged from 77.6% to 87.0%, showing not significant differences. By contrast, PM (30.5%) and CGM (32.3%) presented significantly lower CPD values (P<0.05) as compared with SBC (90.6%). In general, a linear fitting was found between CPD and the release of free amino acids during in vitro protein hydrolysis. The present study provides detailed information, which, combined with the conventional in vitro digestibility studies, may help in the evaluation of different raw sources according to their protein degradation patterns. This information can be applied directly to estimate the protein nutritional quality of ingredients for Senegalese sole feeds.