小鼠H-Y单克隆抗体ELISA检测方法的建立

以纯系 BAL B/ c雄性小鼠脾细胞腹腔注射免疫同系雌性小鼠 9次 ,获得的抗血清经雌、雄鼠脾细胞吸收后用于精子细胞毒性试验 ,测得 H- Y抗血清效价为 1/ 16 0。选取免疫应答最好的雌鼠脾细胞与 SP2 / 0骨髓瘤细胞融合 ,用精子细胞毒性方法筛选效价较高的细胞株制备腹水 ,建立优化的 EL ISA反应条件。优化后的 EL ISA反应条件为 :抗原4℃过夜 ,加 H- Y抗血清 37℃反应 12 0 m in,加 HRP- Ig G 37℃反应 30 min,加 TMB 2 5℃ 30 min。优化后的 EL ISA与常规 EL ISA比较 ,可以明显降低阴性吸光值 ,检测灵敏度从 1/ 32 0上升为 1/ 12 80     

水牛精子在胞质内注射后的早期形态变化

应用胞质内精子注射 (intracytoplasmic sperm injection,ICSI)技术探讨了水牛卵母细胞胞质与注射精子间的相互作用以及 ICSI精子的形态变化。ICSI后 13h,5 9.4 %的精子头部已膨大 ,且有 18.8%的精子进入解聚状态。雌雄原核发育不同步 ,雄原核在 ICSI后 16 h和 19h的形成率分别为 3.2 %和 4 0 .0 % ;雌原核在 ICSI后 13h已达到 71.9% ,16 h时提高到 90 .3%。经离子霉素与 6 - DMAP联合激活 ,ICSI后 19h产生的胚胎核型以 2 PN PB1 为主 ,其雌原核形成 (激活 )率为 91.3% ,雄原核形成率为 4 0 .0 % ,5 1.3%为孤雌胚。用 5 mm ol/ L 的 DTT预处理精子 1h,可提高精子的解聚率 (30 .9%比 12 .7% ,P<0 .0 5 ) ,但对雄原核的形成率无显著影响 (33.3%比 32 .7% ,P>0 .0 5 )。结果表明 ,水牛精子 ICSI后的雄原核形成时间晚于卵子雌原核形成时间 ,且其比率低于卵子 ,DTT处理精子能提高其解聚率 ,但对雄原核形成无影响     

In vitro studies of Norwegian Red bovine semen immobilized and cryopreserved in alginate solid gel network

Development of new semen cryopreservation techniques improving sperm survival and ensuring availability of viable spermatozoa for a prolonged time‐period after AI is promising tools to reduce sensitivity of timing of AI and enhance overall fertility. The SpermVital^® technology utilizes immobilization of bull spermatozoa in a solid network of alginate gel prior to freezing, which will provide a gradual release of spermatozoa after AI. The objective of this study was to compare post‐thaw sperm quality and in vitro sperm survival over time of Norwegian Red bull semen processed by the SpermVital^® (SV) technology, the first commercialized production line of SpermVital^® (C) and by conventional procedure applying Biladyl^® extender (B). Post‐thaw sperm motility was not significantly different between SV, C and B semen (p > .05). However, sperm viability and acrosome intactness were higher for SV than C and B semen (p < .05). Small differences in DNA quality were observed (p < .05). Sperm viability after storage in uterus ex vivo was higher for SV than for C semen (p < .05). Furthermore, sperm survival in vitro over time at physiological temperature was significantly higher for SV semen than C semen as well as B semen during the incubation period of 48 hr (p < .05). In conclusion, the SpermVital^® technology is improved and is more efficient in conserving post‐thaw sperm quality and results in higher sperm viability over time in vitro for SV than for C and B semen.     

Disaccharide combinations and the expression of enolase3 and plasma membrane Ca2+ ATPase isoform in sturgeon sperm cryopreservation

Acipenser sinensis and Acipenser dabryanus are critically endangered species, so germplasm conservation via cryopreservation of sperm is necessary. Disaccharides can act as membrane‐impermeable cryoprotectants, and enolase3 (ENO3) and plasma membrane Ca²⁺ ATPase isoform (PMCA2) are proteins associated with sperm quality. We considered seven characteristics of sperm quality in cultured brood stock from A. sinensis and A. dabryanus. We tested use of sucrose or trehalose alone and in combination at different concentrations for cryopreservation of A. dabryanus sperm. A low concentration of sucrose plus trehalose (S₁₅T₁₅) was optimal. Mixing of the extender with sucrose, lactose, or trehalose alone or with pairwise mixtures revealed that a mixture of lactose and trehalose (L₁₅T₁₅) gave the best results for both A. sinensis and A. dabryanus. Enolase3 and PMCA2 expression levels were measured in cryopreserved A. sinensis sperm via Western blotting. Relative ENO3 and PMCA2 expression levels were examined, and the relationship between disaccharide composition, sperm quality and protein expression was explored in A. sinensis. The results showed that relative ENO3 and PMCA2 expression levels were the highest at L₁₅T₁₅ in cryopreserved A. sinensis sperm. There were significant positive correlations between ENO3 expression and percentage membrane integrity, and between PMCA2 expression and sperm motility parameters (percentage of motile sperm, curvilinear velocity, straight‐line velocity and average path velocity; p < .05) in cryopreserved A. sinensis sperm. Our results indicate the optimal disaccharide combination and concentrations for cryopreservation of A. sinensis and A. dabryanus sperm and suggest that ENO3 and PMCA2 expression levels could serve as a valuable indicator of sperm quality in A. sinensis.     

氮添加对天山高寒草原土壤酶活性和酶化学计量特征的影响

为探究氮添加对高寒草原生态系统土壤酶活性的影响,于2018年在中国科学院巴音布鲁克草原生态系统研究站,选择4个氮添加水平(对照,N0,0 kg·hm^-2·a^-1;低氮,N1,10 kg·hm^-2·a^-1;中氮,N3,30 kg·hm^-2·a^-1;高氮,N9,90 kg·hm^-2·a^-1),开展土壤酶活性对氮添加响应的研究,分析土壤酶活性对氮添加的响应特点,土壤酶化学计量比以及土壤酶活性与土壤环境因子的关系。结果表明:与对照相比,氮添加在N3水平显著增加β-1,4葡萄糖苷酶(βG)、β-D-纤维二糖水解酶(CBH)和β-1,4木糖苷酶(βX)酶活性(P<0.05),N1和N3水平显著增加碱性磷酸酶(AKP)活性(P<0.05),N3水平显著降低多酚氧化酶(PPO)活性(P<0.05),氮添加对亮氨酸氨基肽酶(LAP)活性影响不显著,N3水平下显著增加N-乙酰-β-D氨基葡萄糖苷酶(NAG)活性(P<0.05)。相关分析表明,8种土壤酶活性均与土壤有机碳(SOC、NAG除外)和总磷(TP)显著相关,与土壤总氮(TN)不相关。研究区土壤酶活性C∶N∶P化学计量比为1∶1∶1.2,与全球生态系统的土壤酶活性C∶N∶P的比值1∶1∶1相偏离,表明该研究区土壤微生物生长受磷素限制。冗余分析(RDA)进一步揭示出土壤有机碳和土壤全磷含量是影响土壤酶活性的主要因子。     

基于叶气温差的生育中期冬小麦水分亏缺诊断研究

为了获取叶片尺度基于叶气温差的冬小麦生育中期水分亏缺诊断阈值,分析了冬小麦叶气温差的典型日变化及其与土壤水分、空气温度及太阳辐射等因子的相互关系,揭示了冬小麦叶气温差对光合气体交换参数的影响,确定了冬小麦叶片水分利用效率较适宜的非气孔限制值及叶气温差范围。结果表明,冬小麦叶气温差在不同的土壤水分条件下表现出不同的日变化规律,随光量子通量增加而升高,随土壤水分、气温增加而降低;4、5和6月冬小麦叶片水分利用效率较适宜的非气孔限制值范围分别为0.7~1.3、1.1~2.0和0.9~1.5,叶气温差控制范围分别为-1.2~0.4、-1.5~-1和-1.25~-0.9℃。     

硫酸铜对泥鳅精子活力的影响

以泥鳅精子为受试材料,应用计算机辅助精子分析系统,研究硫酸铜(0.1、1、10 mg/L)对泥鳅的精子活力的影响.结果表明,在0 h试验组,0.1 mg/L的硫酸铜明显降低泥鳅精子运动速度,1 mg/L的硫酸铜对运动时间和速度产生显著抑制作用,10 mg/L的硫酸铜时运动百分数、运动时间和速度都产生明显影响(P<0.0...     

Summer drought decreases Leymus chinensis productivity through constraining the bud,tiller and shoot production

Extreme drought events can directly decrease productivity in perennial grasslands. However, for rhizomatous perennial grasses it remains unknown how drought events influence the belowground bud bank which determines future productivity. Ninety‐day‐long drought events imposed on Leymus chinensis, a rhizomatous perennial grass, caused a 41% decrease in the aboveground biomass and a 28% decrease in belowground biomass. Aboveground biomass decreased due to decrease in both the parent and the daughter shoot biomass. The decreases in daughter shoot biomass were due to reductions in both the shoot number and each individual shoot weight. Most importantly, drought decreased the bud bank density by 56%. In addition, drought induced a bud allocation change that decreased by 41% the proportion of buds that developed into shoots and a 41% increase in the buds that developed into rhizomes. Above results were supported by our field experiment with watering treatments. Thus, a 90‐day‐long summer drought event decreases not only current productivity but also future productivity, because the drought reduces the absolute bud number. However, plasticity in plant development does partly compensate for this reduction in bud number by increasing bud development into rhizomes, which increases the relative allocation of buds into future shoots, at the cost of a decrease in current shoots.     

Evaluation of direct and indirect phosphorus limitation of methanogenic pathways in a calcareous subtropical wetland soil

The effect of phosphorus (P) and carbon (C) on methanogenesis was investigated in a low-P (130 mg P kg⁻¹ soil) wetland within Everglades National Park. Soil was amended with C substrates (acetate, formate, butyrate, and glucose) with or without P, and CO₂ and CH₄ production was monitored. Production of CH₄ increased with P addition although no effect on CO₂ was observed. Methane production was stimulated by all C substrates except for butyrate. No effect of C on CO₂ production was observed except for stimulation following glucose addition. Production of CH₄ following formate addition was not affected by P, suggesting hydrogenotrophic methanogens may be substrate, not P, limited. Addition of P to all other C substrates heightened CH₄ production and lowered the CO₂–C:CH₄–C ratio relative to the corresponding C only treatment, suggesting that P may have limited acetoclastic methanogens and fermentation.