鸡白细胞介素2原核表达载体的构建及表达产物的鉴定

将鸡白细胞介素2(IL-2)的cDNA克隆到原核表达载体pET-30a( )上，构建原核重组表达质粒pET-IL-2，并在大肠杆菌BL21(DE3)中诱导表达，37℃用IPTG诱导3.5h后，SDS-PAGE检测表明，表达蛋白以包涵体的形式存在。将诱导后的工程菌超声波裂解，离心后的沉淀用PBS洗涤5～6次，得到高纯度的目的蛋白。     

镉对小鼠卵巢颗粒细胞凋亡的影响研究

为探讨镉对雌性生殖机能的影响,幼鼠皮下注射孕马血清促性腺激素(PMSG),48 h后腹腔注射不同剂量的氯化镉,用琼脂糖凝胶电泳及TUNEL法对卵巢颗粒细胞进行凋亡检测;免疫组织化学法检测颗粒细胞的bax表达。结果显示注射氯化镉后48 h的高、中、低剂量组卵巢颗粒细胞(GC)凋亡率分别为(13.887 1±1.329 9)%、(10.757 3±1.229 3)%(、10.020 8±0.983 7)%,均显著高于对照组(0.637 8±0.387 2)%(P<0.01);96 h对照组GC凋亡率显著升高,高剂量组与其它组间差异明显,中、低剂量组与对照组差异不明显。凝胶电泳仅96 h高剂量组呈现梯状带。48 h、96 h各剂量组bax表达明显高于对照组(P<0.01)。可见镉对卵巢颗粒细胞的凋亡有促进作用,并有一定的剂量相关性;TUNEL法检测细胞凋亡优于电泳法,当细胞凋亡率较低时(<20%),电泳检测不到DNA梯状带;卵巢颗粒细胞凋亡和bax的表达有一定的相关性,随凋亡率的升高bax表达呈上升趋势。     

Application of time-resolved fluorometry to immunoassays for bovine reproductive hormones

The principle of time‐resolved fluorometry with lanthanide chelates was established in the 1980s, but in the field of animal sciences it has not been widely applied to immunoassays. However, immunoassays that utilize time‐resolved fluorometry are possible alternatives to radioimmunoassays, since they can attain high sensitivity without safety risks. In this short review, we introduce the development of time‐resolved immunoassays for inhibin A, inhibin B and follicle‐stimulating hormone (FSH), and describe their application to the investigation of FSH regulation in male and female cattle. The results obtained using these newly developed immunoassays indicate that inhibin A acts as a feedback regulator for FSH secretion in female cattle, whereas inhibin A, and probably inhibin B, do so in male cattle.     

半胱亚磺酸脱羧酶(CSD)基因在大鼠睾丸组织中的表达及其序列分析

试验根据大鼠肝脏牛磺酸生物合成关键酶——半胱亚磺酸脱羧酶(CSD)基因已知核苷酸序列,设计了1对特异的寡核苷酸引物,对提取的睾丸组织总RNA进行RT-PCR扩增,扩增出468 bp的DNA片段,在国际上首次证明CSD基因在大鼠睾丸组织中存在表达,即牛磺酸可在大鼠睾丸组织中生物合成。测定了扩增的睾丸组织CSD基因核苷酸序列,并与已知肝脏CSD基因核苷酸序列及其对应的氨基酸序列作了同源性分析。结果表明:睾丸组织CSD基因核苷酸序列同已知肝脏CSD基因核苷酸序列的同源性为99.8%,其对应氨基酸序列的同源性为100%。     

抽提条件对酶制剂中果胶酶活力测定的影响

用0．1mol/L氯化钠、0．02mol/L,pH7．4的磷酸缓冲液和蒸馏水,于40℃和4℃对浙江酶、华芬酶和Avizyme－1500酶制剂分别抽提1．5小时和12小时,对3种酶制剂中的果胶酶活力进行测定。结果表明：用0．1mol／L氯化钠于4℃抽提12小时,对浙江酶制剂中的果胶欧抽提效果最好;用蒸馏水于4℃抽提12小时,对华芬酶和Avizyme－1500酶制剂中的果胶酶抽提效果最好。     

ARF基因干扰表达对不同发育阶段和贮藏条件马铃薯酶活性的影响

以前期研究获得的马铃薯栽培品种“甘农薯2号”ADP核糖基化因子(ARF)基因干扰表达转化植株为材料,用qRT-PCR法对转基因植株的ARF基因表达量进行了测定,结果表明,不同发育阶段转基因叶片中ARF基因的相对表达量先降低,生长后期略有增加,表明ARF基因干扰表达量随马铃薯生长发育发生改变。ARF基因干扰表达影响马铃薯叶片中酶活性,不同发育阶段转基因叶片与对照相比,多酚氧化酶(polyphenol oxidase, PPO)活性升高11.61%~27.84%,硝酸还原酶(nitrate reductase, NR)活性提高21.10%~41.32%,磷脂酶D(phospholipase D, PLD)活性降低2.88%~57.64%,蔗糖磷酸合成酶(sucrose phosphate synthase, SPS)活性提高29.00%~39.57%;不同温度(4℃和室温)贮藏的块茎中ARF基因的相对表达量变化趋势一致:均先降低,再升高,但前者较后者ARF基因相对表达量显著降低。室温较4℃贮藏的转基因块茎PPO活性升高30.44%~56.28%,NR活性提高17.41%~40.92%,PLD活性降低24.39%~85.11%,SPS活性提高30.89%~45.78%。室温较4℃贮藏的非转基因块茎PPO活性升高25.11%~70.66%,NR活性提高36.07%~89.62%,PLD活性降低11.35%~72.64%,SPS活性提高27.31%~61.33%。本研究通过探讨ARF基因干扰表达对马铃薯生理生化特性的影响,为进一步研究ARF基因在马铃薯生长发育调控中的作用提供一定的理论基础。     

Seasonal and Diurnal Variation in Water-Soluble Carbohydrate Concentrations of Repeatedly Defoliated Red and White Clovers in Central Kentucky

Nonstructural carbohydrates of pasture plants, comprising water-soluble carbohydrates (WSCs) and starch, may contribute to excessive consumption of rapidly fermentable carbohydrates by grazing horses. Seasonal and diurnal variation in WSCs were studied in red (Trifolium pratense L.) and white clovers (Trifolium repens L.) subjected to a typical management regime of rotationally grazed horse pastures. Two red and two white clover cultivars from monoculture plots were harvested after 4 weeks of growth from April to October of 2015, in the morning and afternoon of each harvest date. Water-soluble carbohydrates were quantified for each harvest, and starch was quantified for two harvests. Mean monthly WSC concentrations ranged from 80 to 99 mg/g (freeze-dried weight basis), whereas mean starch concentrations were 31 and 40 mg/g. In September, white clover had 14% more WSCs than red clover (P < .0001). Water-soluble carbohydrate concentrations were 10% higher in the afternoon than in the morning (P < .0001). Starch concentrations were 290% higher in the afternoon than in the morning (P < .0001), and nonstructural carbohydrate concentrations in the afternoon averaged 150 mg/g. Further studies are needed to determine whether the mixed grass-legume pastures of central Kentucky accumulate enough nonstructural carbohydrates to present risk factors for equine metabolic or digestive dysfunction.     

钙依赖蛋白激酶基因表达载体的构建及其对拟南芥的遗传转化

将钙依赖蛋白激酶(CDPK)基因插入到植物表达载体pBI121上,构建成特异性表达载体pBI121-CDPK,并采用农杆菌介导法将其转化进入拟南芥Arabidopsis thaliana Columbia生态型品种中,获得了卡那霉素筛选的抗性植株,通过PCR扩增验证,拟南芥基因组中含有此抗性基因.     

抗阿维菌素抗体的研制

本文合成了二个模拟阿维菌素特征结构的半抗原，４‘＝Ｏ－（单）琥珀酰阿维菌素和５－Ｏ－（单）琥珀酰阿维菌素。将半抗原与载体蛋白连接，制成人工抗原。