17α-甲基睾酮对稀有鮈鲫精巢DNA甲基转移酶基因表达的影响

[目的] 以稀有鮈鲫雄鱼为研究对象,探讨17α-甲基睾酮（17α-methyltestosterone,MT）对稀有鮈鲫精巢甲基转移酶（DNMT）基因相对表达量的影响。[方法] 采用不同浓度MT（25、50和100 ng/L）处理稀有鮈鲫雄鱼7、14和21 d,处理结束时,每组随机解剖10条鱼,取精巢组织,Trizol一步法提取总RNA,实时荧光定量PCR（qRT-PCR）检测DNA甲基转移酶基因（dnmts）相对表达量。[结果] MT处理稀有鮈鲫雄鱼7 d,25 ng/L MT处理组（P<0.05）、50 ng/L MT处理组（P<0.01）和100 ng/L MT处理组（P<0.05）精巢dnmt3基因相对表达量显著（或极显著）降低;25 ng/L MT处理组和50 ng/L MT处理组精巢dnmt5基因相对表达量显著（P<0.05）降低;50 ng/L MT处理组精巢dnmt7基因相对表达量显著（P<0.05）降低。MT处理稀有鮈鲫雄鱼14 d,100 ng/L的MT处理组dnmt3、dnmt5、dnmt6、dnmt7基因相对表达量显著（P<0.05）升高;50 ng/L的MT处理组dnmt1基因相对表达量极显著（P<0.01）升高。MT处理稀有鮈鲫雄鱼21 d,50 ng/L MT处理组和100 ng/L MT处理组dnmt1基因相对表达量显著（P<0.05）升高;100 ng/L MT处理组dnmt8基因相对表达量极显著（P<0.01）升高。[结论] MT可以干扰稀有鮈鲫精巢DNA甲基转移酶基因dmmts的mRNA表达。MT对DNMTs酶活性的影响以及是否可以通过改变DNA甲基化水平对稀有鮈鲫精子的发生产生影响有待研究。     

The Effect of Levels of α-Galactosidase Enzyme on Performance of Broilers Fed Diets Based on Corn and Soybean Meal

A study was conducted with male chicks of a commercial broiler strain to evaluate the effects of different dosage levels of a commercial α-galactosidase enzyme. Diets were formulated based on corn and soybean meal to meet the nutrient standards of top poultry companies. The positive control diet was formulated with no adjustment in the ME content of the soybean meal associated with enzyme supplementation. The negative control diet was formulated assuming a 10% improvement in the ME of the soybean meal. The negative control diet was supplemented with 0, 1.5, 3.0, 4.5, or 6.0 g of enzyme per kilogram of soybean meal to provide for 0, 45, 90, 135, or 180 galactosidase units (GALU)/kg of soybean meal. Each of the test diets was fed in mash form to 8 replicate pens of 30 birds. Body weight, feed efficiency, mortality, and calorie conversion were determined at 14, 35, and 42 d of age. Body weight, feed efficiency, and mortality were not significantly (P < 0.05) affected by dietary treatment. Birds fed the negative control with no enzyme supplement were numerically reduced in body weight or feed efficiency as compared with the control diet; however, addition of the enzyme was without benefit. When unadjusted energy values were analyzed, birds fed the negative control diet were more efficient in calorie conversion, but this finding was unrelated to enzyme addition, suggesting that diet composition was responsible for the differences in calorie utilization. When adjusted energy values were compared, birds fed the negative control tended to have higher (less efficient) calorie conversion than those fed the positive control with little or no indication of improvement from the addition of the α-galactosidase enzyme. Results of this study show no benefit from the addition of the enzyme used in this trial.     

饲用α-半乳糖苷酶酶活稳定性的研究

在酶制剂生产加工和储存过程中,酶活稳定性是酶应用过程中最常遇到的实际问题。本文研究不同载体、高温高湿、金属离子、保存初始pH值、保护剂浓度和存储时间对体外α-半乳糖苷酶稳定性的影响。试验结果表明:①无机载体碳酸钙、元明粉和滑石粉对α-半乳糖苷酶活性影响很大,收率均为0%;有机载体中玉米皮粉效果最好,收率高达90.83%;其次是淀粉,收率为61.11%;稻壳粉收率最低,仅为44.72%。②湿度为17%,温度小于85℃,酶活损失率小于10%,说明该酶有较强的耐温能力,温度为90℃,酶活损失率达到16.2%,随着温度上升,酶活损失率开始增加,不利于酶活的保存。③Cu2+对该酶有抑制作用3,7℃恒温水浴4 h后酶活存留率仅为74%,Mn2+、Zn2+、Ca2+、Na+、K+和Mg2+对α-半乳糖苷酶有几近相同的保护作用,酶活存留率均大于92%。④该酶最适保存pH值为5.3;pH值为4.4时,酶活损失率达到30.1%,不利于酶的保存;pH值为5.6时,酶活损失率为3.77%,pH值在4.7~5.6之间,酶活损失率小于10%,有利于酶的保存。总之,过酸和过碱都不利于酶的保存。⑤1%NaCl、1%甘露醇、5%和7%的山梨醇都有杂菌产生,不利于酶的保存3;%~5%的NaCl酶活损失小于10%1,%和5%~7%的甘油酶活损失率小于5%,都有利于酶活保存。     

α-硫辛酸对热应激绵羊生长性能、抗氧化及免疫的影响

本试验旨在研究α-硫辛酸(α-LA)对热应激绵羊生长性能、抗氧化及免疫的影响,以期阐述其抗热应激的机理。选择4～5月龄、体重35～40 kg的绵羊15只,随机分为对照组(CTL)、低剂量组(LA-L,添加600 mg/kgα-LA)、高剂量组(LA-H,添加900 mg/kgα-LA),在热应激条件下(7～8月)饲喂40 d。结果表明:1)与对照组相比,α-LA添加组平均日采食量增加,低剂量组和高剂量组分别比对照组增加了7. 3%和2. 7%(P> 0. 05);高剂量组血清中丙氨酸转氨酶(ALT)、门冬氨酸转氨酶(AST)活性及α-LA添加组血浆中丙二醛(MDA)、肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、白细胞介素-8(IL-8)含量显著下降(P<0.05);α-LA添加组血浆中三碘甲腺原氨酸(T3)、热休克蛋白70(HSP70)、免疫球蛋白G(IgG)含量及超氧化物歧化酶(SOD)活性显著升高(P <0.05)。2)与对照组相比,α-LA添加组肝脏中炎性相关因子IL-1β、IL-8、核因子-κB(NF-κB) mRNA表达量显著下降(P <0.05),抗炎因子干扰素-γ(IFN-γ)mRNA表达量显著升高(P<0.05)。综上,α-LA显著增强热应激绵羊的抗氧化能力,减少炎症相关因子的表达,增强抗炎因子的表达,提高机体免疫机能,且与α-LA剂量有关。     

Cloning,Expression of Clostridium perfringens α-toxin Gene and Preparation of its Antisera

One pair of primers had been designed and synthesized based on the α-toxin gene of Clostridium perfringens.The complete α-toxin gene fragment was amplified by polymerase chain reaction (PCR), and then was cloned into pGEM-T Easy vector to construct pGEM-T-α.Digested with EcoRⅠ and Hind Ⅲ, a fragment of 1125 bp was cloned into the expression plasmid vector pET-28a(+).The recombinant plasmid was transformed into the BL21(DE3)plys and induced by 1.0 mmol/L IPTG at 37 ℃.The expression product was found to be 46.1 ku as expected one identified by SDS-PAGE, and confirmed by Western blotting with Clostridium perfringens type A antisera, indicating similar reactivity with native α-toxin.Recombinant α-toxin protein was simultaneously found in culture supernatant, postsonic supertanant and inclusion bodies, most protein was expressed in inclusion bodies, which indicated recombinant α-toxin protein was expressed in the extracellular, periplasm and cytoplasm.Recombinant α-toxin protein in postsonic supertanant could not make mice die, indicating its non-toxicity.Toxin-antitoxin neutralization test showed that antisera of recombinant α-toxin protein were specific to α-toxin.Upon immunization of rabbit with the recombinant α-toxin protein, antisera with high antibody titer neutralizing 100 MLD toxin per 1 mL were prepared.     

添加黑麦草饲粮对鹅肉脂肪酸组成的影响

本研究旨在观察饲粮中添加不同比例多花黑麦草(Lolium multiflorum Lam)对鹅肉脂肪酸组成影响,揭示牧草摄食量与鹅肉α-亚麻酸(ALA)相对含量之间的关系。选用50羽14日龄长乐灰鹅,分成5个处理,饲养期60 d。各处理饲粮中黑麦草干物质比例分别为: 0.00%(CK),5.42%,11.92%,19.52%和27.24%。结果表明:随着饲粮中黑麦草比例的提高,鹅肉ALA相对含量显著增加,试验各组分别比对照组增加了86.90%,197.24%(P<0.05),393.10%(P<0.05)和304.83%(P<0.05);PUFA/SFA显著上升(P<0.05);n-6PUFA/n-3PUFA显著下降(P<0.05)。鹅肉ALA相对含量与饲粮中黑麦草比例的曲线回归模型方程为:y=0.270+0.106x-0.002x²(F=22.99^**)。当饲粮中黑麦草的干物质比例在0%～26.50%时,鹅肉ALA的含量呈上升趋势;当黑麦草的比例超过26.50%时,鹅肉ALA的含量转而呈下降趋势。说明黑麦草中的ALA能够通过摄食消化过程沉积到鹅肉中。     

Cloning and Protein Structure Analysis of Hypoxia Inducible Factor-1α Gene in Yak

In this study,hypoxia inducible factor-1α(HIF-1α) gene was cloned from yak spleen by RT-PCR,and it was divided into two sections for amplification.The sequence and protein structure were analyzed using related bioinformatics tools.The length of the coding region was 2 472 bp,which encoded 823 amino acids.Its molecule mass was 92.13 ku,and PI was 5.09.It displayed as a hydrophilic protein,whose secondary structure was mainly composed of random coil and α-helix.It had 69 phosphorylation sites and 4 N-glycosylation sites.Phylogenetic tree displayed that Maiwa yak,Bos grunniens,Bos mutus and Bos taurus gathered into one cluster.Maiwa yak HIF-1α gene was successfully cloned in this study,it provided a viable reference for further study of genetic characteristics of HIF-1α.     

低分子质量壳寡糖对蛋鸡生产性能、蛋品质、血清生化指标、盲肠微生物数量及脾脏白细胞介素-2、肿瘤坏死因子-α基因表达的影响

本试验旨在研究饲粮中添加不同水平的低分子质量(1 000 u)壳寡糖对蛋鸡生产性能、蛋品质、血清生化指标、盲肠微生物数量以及脾脏白细胞介素-2(IL-2)和肿瘤坏死因子-α(TNF-α)基因表达的影响。选取体重和产蛋率相近的58周龄海兰褐壳蛋鸡600只,随机分为4组,每组5个重复,每个重复30只。对照组饲喂基础饲粮,试验组分别饲喂在基础饲粮中添加300、600、900 mg/kg壳寡糖的试验饲粮。预试期为7 d,正试期为42 d。结果表明:1)添加300、600和900 mg/kg壳寡糖组的产蛋率分别比对照组提高了4.52%(P0.05)、2.99%(P0.05)和4.08%(P0.05)。2)试验第3、6周末,添加600和900 mg/kg壳寡糖组的鸡蛋哈夫单位分别比对照组提高了6.87%、6.69%和6.47%、6.60%(P0.05)。3)与对照组相比,饲粮添加600、900 mg/kg壳寡糖显著降低了血清葡萄糖、胆固醇含量和谷草转氨酶活性(P0.05)。4)与对照组相比,饲粮添加600、900 mg/kg壳寡糖显著提高了盲肠双歧杆菌和乳酸杆菌的数量(P0.05),显著降低了盲肠金黄色葡萄球菌的数量(P0.05)。5)与对照组相比,饲粮添加300、600 mg/kg壳寡糖显著提高了脾脏IL-2 m RNA表达水平(P0.05),饲粮添加600 mg/kg壳寡糖显著提高了脾脏TNF-αm RNA表达水平(P0.05)。由此可见,饲粮中添加不同水平的壳寡糖,提高了蛋鸡的产蛋率和哈夫单位,调节了肠道微生物菌群,增强了蛋鸡的免疫力,适宜壳寡糖添加水平为600 mg/kg。     

α-酮戊二酸和大蒜素对生长猪生长发育及养分表观消化率的影响

本试验旨在研究α-酮戊二酸(AKG)和大蒜素对生长猪生长性能、养分表观消化率、血清生化指标以及肠道形态结构的影响。选用40头初始平均体重为(30.64±1.35)kg的健康三元(杜×长×大)杂交生长猪,按体重相近、性别比例相同原则随机分为5组,每组8个重复,每个重复1头猪。5组试验猪分别饲喂无抗基础饲粮(对照组)、无抗基础饲粮+25 mg/kg抗生素(Ⅰ组)、无抗基础饲粮+1%AKG(Ⅱ组)、无抗基础饲粮+0.5%大蒜素(Ⅲ组)、无抗基础饲粮+1%AKG+0.5%大蒜素(Ⅳ组)。预试期7d,正试期21d。结果表明:1)与对照组相比,Ⅰ组和Ⅳ组生长猪的平均日增重分别提高了10.80%和9.91%,但差异不显著(P0.05)。2)与对照组相比,Ⅱ组钙的表观消化率显著提高(P0.05),Ⅳ组粗蛋白质、钙和磷的表观消化率显著提高(P0.05),且Ⅳ组较Ⅰ组显著提高了磷的表观消化率(P0.05)。3)与对照组相比,Ⅱ组血清免疫球蛋白G(IgG)含量显著提高(P0.05);Ⅲ组血清甘油三酯(TG)含量显著降低(P0.05);Ⅳ组血清总胆固醇(TCHO)和TG含量显著降低(P0.05),血清碱性磷酸酶(AKP)、谷草转氨酶(AST)和谷丙转氨酶(ALT)活性及血清免疫球蛋白A(IgA)、IgG含量显著提高(P0.05)。与Ⅰ组相比,Ⅳ组血清TCHO和TG含量有降低趋势(P0.05),血清AST和ALT活性显著提高(P0.05)。4)与对照组相比,Ⅳ组回肠绒毛高度及空肠和回肠的绒毛高度/隐窝深度值显著提高(P0.05)。5)与对照组相比,Ⅳ组回肠中杯状细胞和淋巴细胞数目显著增加(P0.05)。综上,无抗饲粮中联合添加1%AKG和0.5%大蒜素可通过改善肠道形态结构,提高对粗蛋白质、钙和磷等养分的消化吸收能力,促进肠道健康,进而改善生长猪的生长性能。