海藻糖对牛未成熟卵母细胞玻璃化冷冻的影响

【目的】研究胞外冷冻保护剂海藻糖对玻璃化冷冻-解冻后牛未成熟卵母细胞形态、活性线粒体分布和核成熟的影响。【方法】将未成熟卵母细胞随机分为新鲜组、不同浓度海藻糖(0.25、0.5和1 mol/L)组和0.5 mol/L蔗糖组共5组。新鲜组作为玻璃化冷冻的对照组,海藻糖和蔗糖组均采用两步法进行玻璃化冷冻和解冻处理。在冷冻的第1步不添加海藻糖和蔗糖,在冷冻的第2步和解冻的第1步分别添加对应浓度的海藻糖和蔗糖,在解冻的第2步海藻糖和蔗糖浓度均减半。解冻后,统计卵母细胞形态正常率;用JC-1染色检测卵母细胞的活性线粒体分布区域和荧光强度;体外成熟培养24 h后,统计卵母细胞的核成熟率。【结果】玻璃化冷冻组卵母细胞的形态正常率和核成熟率均显著低于新鲜组卵母细胞(P<0.05),0.5 mol/L海藻糖组卵母细胞形态正常率和核成熟率均显著高于0.5 mol/L蔗糖组及0.25和1 mol/L海藻糖组(P<0.05);0.5 mol/L海藻糖组卵母细胞的活性线粒体均匀分布在细胞膜内侧区域,在绿光和蓝光下分别呈现强的红色荧光和黄绿色荧光,且荧光强度高于其他玻璃化冷冻组卵母细胞,但仍低于新鲜组...     

海藻糖替代部分甘油对延边黄牛冻融精子质量的影响

【目的】探讨是否可以通过补充海藻糖来降低冷冻保护剂中甘油的浓度,从而提高冻融精子的质量。【方法】分别用6%甘油(Ⅰ组)及3%甘油+0、50、100和150 mmol/L海藻糖(Ⅱ、Ⅲ、Ⅳ和Ⅴ组)处理精子,用精子-微生物动(静)态图像检测分析系统(CASA)检测冻融精子的活力、质膜完整性和顶体完整性及动力学相关参数,筛选出最佳海藻糖处理浓度用于后续试验。通过Western blotting方法检测精子蛋白酪氨酸磷酸化水平评价精子获能状态,Hoechst 33342/PI/JC-1联合染色法检测精子的活率及线粒体膜电位,色霉素A₃(CMA₃)染色法检测精子DNA的完整性,部花青540(M540)和Yo-Pro-1染色检测精子膜脂质紊乱水平。【结果】与Ⅰ组相比,Ⅱ组精子的活力、质膜完整性、顶体完整性以及曲线速度(VCL)和直线率(STR)均显著下降(P<0.05),Ⅲ、Ⅳ和Ⅴ组组精子的活力、VCL、直线速度(VSL)、平均路径速度(VAP)、直线性运动(LIN)和STR均显著升高(P<0.05)。因此,后续试验选用100 mmol/L海...     

大肠杆菌海藻糖磷酸合酶基因的克隆

根据报道的大肠杆菌（Escherichia coli)海藻糖－6－磷酸合酶基因（otsA)，设计引物，通过PCR技术从大肠杆菌XLI菌株的总DNA中扩增到一个1．4kb片段。经克隆、测序分析，该片段长1425bp并含一完整的开放读框（ORF）。在核苷酸水平上，该ORF与已报道的otsA基因具有99．86％的同源性。在氨基酸水平上，其推断性的编码产物蛋白与OtsA具有100％一致性。     

金龟子绿僵菌诱发UV-B抗性导致分生孢子产孢和孢子逆境耐受性的转化

The conidial tolerance of Metarhizium anisopliae var.anisopliae isolate ARSEF 2575 to UV-B irradiation is greatly influenced by growth-environment alterations. In this review, we report high variability in conidial UV-B tolerance in response to altered culture conditions.Conidia produced on insect cadavers[Zophobas morio(Coleoptera) or Galleria mellonella(Lepidoptera)] had low tolerance to UV-B radiation; and conidia produced on potato dextrose agar supplemented with yeast extract (PDAY) had medium UV-B tolerance; whereas conidia produced on a minimal medium without any carbon source (MM), on MM with a non-preferred carbon source such as lactose (=MML), on PDAY plus 1 M NaCl or KCl, or PDBY with high alkalinity had the highest UV-B tolerances. All of the above conditions that induced high UV-B tolerance, however, also greatly reduced conidial production. Comparisons between stress tolerance and conidial production, particularly with conidia produced under osmotic and nutritive stress, point out that the benefits of producing very tolerant conidia have the enormous cost of low conidial production. Growth under visible light also greatly improved conidial UV-B tolerance, but light did not negatively influence conidial production. Therefore, culture on rich media under light is proposed as the most promising approach to producing conidia with improved UV-B tolerance for biological control of pest insects in agriculture.     

Transformation of tobacco with an Arabidopsis thaliana gene involved in trehalose biosynthesis increases tolerance to several abiotic stresses

Summary Trehalose (a non-reducing disaccharide) plays an important role in abiotic stress protection. It has been shown that using trehalose synthesis genes of bacterial origin, drought and salt tolerance could be achieved in several plants. A cassette harboring the AtTPS1 gene under the control of the CaMV35S promoter and the Bialaphos resistance gene was inserted in the binary plasmid vector pGreen0229 and used for Agrobacterium-mediated transformation of tobacco (Nicotiana tabacum). T0 plants obtained were analyzed by PCR for the presence of AtTPS1 gene. Thirty lines were positive and seeds were germinated on media with 6 mg/l PPT to obtain T1 plants that were grown in the greenhouse to obtain T2 seeds that were germinated on selective media. Lines which seeds showed a 100 % survival rate were considered homozygous transgenic T1 lines. Three lines were selected and gene expression confirmed by northern and western blots. Transgenic seeds were germinated on media with different concentrations of mannitol (0, 0.25, 0.5 and 0.75 M) and sodium chloride (0, 0.07, 0.14, 0.2, 0.27 and 0.34 M) to score their tolerance to osmotic stress. Assays were conducted to test the tolerance of transgenic plants to drought (measurement of water percentage as a consequence of water withdrawal), desiccation (measurement of water loss as a consequence leaf detaching) and temperature stresses (germination at 15 ^∘C and 35^∘C). Transgenic tobacco plant lines registered higher germination rates under osmotic and temperature stress situations than did wild-type plants. Responses to drought and desiccation stresses were similar for all plant lines. It can hence be suggested that the heterologous expression of TPS1 gene from Arabidopsis can be used successfully to increase abiotic stress tolerance in model plants and probably in other crops.     

转CrylAC及CrylAC+CpTI基因对棉花上绿盲蝽2种消化酶活性及海藻糖含量的影响

 以转基因棉国抗22,sGK321及其亲本棉泗棉3号、石远321为材料,研究了上述4种寄主上绿盲蝽淀粉酶、蛋白酶活性及海藻糖含量。结果表明：与亲本棉相比,2种转基因棉绿盲蝽若、成虫淀粉酶、蛋白酶活性和海藻糖含量均值较高,且若虫淀粉酶活性、成虫海藻糖含量在转基因棉国抗22和其亲本棉泗棉3号之间差异显著;同一品种棉花上成虫的淀粉酶、蛋白酶活性和海藻糖含量均高于若虫,并差异极显著。绿盲蝽若、成虫淀粉酶、蛋白酶活性及海藻糖含量在转基因棉及其亲本棉上的频率分布高峰基本一致;但与亲本棉花相比,2种转基因棉所测指标在低区间段分布较少,在高区间段分布较多。     

高效阴离子色谱-脉冲安培检测法分析食用菌中海藻糖、甘露醇和阿糖醇

采用高效阴离子色谱-脉冲安培检测法(HAPEC-PAD)测定17种食用菌中海藻糖、甘露醇和阿糖醇含量.结果表明:阿糖醇、海藻糖和甘露醇的线性范围分别为239.62～1.25、59.41～0.15、148.29～5.00μg/mL,检出限分别为0.41、0.04、.11 μg/mL,定量限分别为1.14、0.10和5.79μg/mL;17种食用菌中,香菇(Lentinula edodes)和猴头菌(Hericium erinaceus)中阿糖醇含量较高,茶树菇(Agrocybe aegerita)、刺芹侧耳(Pleurotus eryngii)、鲍鱼侧耳(Pleurotus abalonus)、松口蘑(Tricholoma matsutake)、蛹虫草(Cordyceps militaris)中海藻糖含量较高,姬松茸(Agaricus blazei)、鸡油菌(Cantharellus cibarius)、金顶侧耳(Pleurotus citrinopileatus)、冬虫夏草(Cordyceps sinensis)中甘露醇含量较高;该项检测技术分离效果好、样品测定时不需要衍生化处理、检测时间短,是测定食用菌中功能性寡糖和糖醇类物质的有效方法.     

Different effects of ionic and non-ionic compounds on the freeze denaturation of myofibrils and myosin subfragment-1

The effects of non-ionic (sorbitol, maltose, trehalose) and ionic compounds (Na-glutamate, Na-acetate, Na-sulfate, ammonium sulfate) on freeze denaturation of myosin subfragment-1 (S-1) and of myofibrils were compared. Sugars, Na-glutamate and Na-acetate well suppressed the freeze denaturation of myofibrils as well as S-1 in a concentration dependent manner. Although sulfate suppressed freeze denaturation of S-1 irregularly, it accelerated myofibril denaturation. It was concluded that sulfate salts were useless as cryoprotectant for myofibrils. Stabilization extent by F-actin in frozen storage was much less than that in heating.     

不同酶标抗体稀释液对酶标抗体稳定性的影响

为筛选一种较好的酶标抗体稀释液,试验以磷酸盐缓冲液（PBST）为基础溶液,通过添加甘油、酪蛋白、海藻糖及防腐剂Proclin 300等试剂配制了4种不同组合的酶标抗体稀释液,分别在4℃和37℃条件下考察其对酶标抗体稳定性的影响。结果表明,处理5（0.01 mol/L PBST＋3%酶稳定剂M＋3%海藻糖＋0.05%Proclin 300）的保护效果最好,在4℃保存6个月,酶标抗体的活性仅降低18%。