重组大肠杆菌E.coli P84A/MC1061发酵生产卤醇脱卤酶的研究

用基因工程大肠杆菌E.coliP84A/MC1061生产卤醇脱卤酶,此后,进一步进行了酵母浸出物FM888与酵母蛋白胨FP101的联合调控作用研究。结果表明,用高溶解度酵母浸出物FM888替代原配方中的牛骨蛋白胨,可使E.coliP84A/MC1061生长速率增大,发酵稳定期的菌体浓度保持在较高水平,酶活提高了约2.4倍。当补料培养基中的FM888与酵母蛋白胨FP101的比例为7∶3时,菌体浓度保持在较高水平(OD600≥120),酶活进一步提高约50%,大大提高了单罐产率。此研究可为其他酶制剂的发酵生产提供借鉴。     

酵母细胞壁多糖与铝硅酸盐复合物对猪生长性能、免疫指标及养分消化率的影响

本研究旨在探讨酵母细胞壁多糖(yeast polysaccharide,YP)与铝硅酸盐复合物(alumi-nosilicate complex,AC)对猪的生长性能、免疫指标及养分消化率的影响。试验选用平均体重为(66.18±1.26)kg的"杜×长×大"三元杂交猪140头,随机分成5组,每组4个重复,每个重复7头猪。各组分别饲喂添加不同剂量复合物的试验饲粮,A组为对照组,不添加复合物;B组添加0.1%YP;C组添加0.1%YP和0.9%AC;D组添加0.1%YP和1.9%AC;E组添加0.1%YP和2.9%AC。试验期51 d。结果表明:1)D、E组平均日增重显著低于对照组(P<0.05),各组平均日采食量和料重比差异不显著(P>0.05);2)与对照组相比,D组血清谷胱甘肽过氧化物酶(GSH-Px)活性及补体3(C3)、免疫球蛋白M(IgM)、免疫球蛋白A(IgA)、免疫球蛋白G(IgG)水平显著提高(P<0.05),血清超氧化物歧化酶(SOD)活性和补体4(C4)水平无显著差异(P>0.05);3)各组养分消化率无显著差异(P>0.05)。由此可见,YP和AC复合使用降低了育肥猪的生长性能,0.1%YP和1.9%AC同时添加可增强生长猪的免疫功能。     

酵母壁多糖对断奶仔猪肠道挥发性脂肪酸和微生物菌群的影响

本研究旨在探讨饲粮中添加酵母壁多糖对断奶仔猪肠道挥发性脂肪酸和微生物菌群的影响。试验采用单因素试验设计方法,选取180头遗传背景一致、健康状况良好、胎次和体重接近的21日龄断奶仔猪,随机分为4个组,每组5个重复,每个重复9头猪。4个组试验猪分别饲喂对照饲粮(未添加酵母壁多糖)、0.15%酵母壁多糖饲粮、0.30%酵母壁多糖饲粮和0.45%酵母壁多糖饲粮。试验期21 d。结果表明:1)与对照组相比,饲粮中添加0.15%、0.30%和0.45%酵母壁多糖显著提高了仔猪结肠乙酸的含量(P0.05);其中,0.30%和0.45%酵母壁多糖还显著提高了结肠丙酸、丁酸以及总挥发性脂肪酸的含量(P0.05),且二者之间差异不显著(P0.05)。2)与对照组相比,饲粮中添加0.15%、0.30%和0.45%酵母壁多糖显著降低了盲肠沙门氏菌和大肠杆菌数量(P0.05),且0.30%和0.45%酵母壁多糖组之间差异不显著(P0.05)。由此可知,酵母壁多糖可提高仔猪肠道挥发性脂肪酸含量,并改善肠道微生物菌群结构,根据回归方程预测,酵母壁多糖在仔猪饲粮中的适宜添加水平为0.31%~0.40%。     

Screening and Analysis of Proteins Interacting with TaPDK from Physiological Male Sterility Induced by CHA in Wheat

To further research the regulatory network of pyruvate dehydrogenase kinase (designated as TaPDK) in physiological male-sterility (PHYMS) of wheat induced by chemical hybridizing agent (CHA) SQ-1, an anther cDNA library was constructed, and the proteins interacting with TaPDK were screened via yeast two-hybrid technique. Subsequently, a few candidate proteins in nucleotide expression levels were detected by real-time quantitative PCR. Yeast-two hybrid screening was performed by mating yeast strain Y2HGold containing BD-TaPDK bait plasmid with yeast strain Y187 including anther cDNA library plasmid. Diploid yeast cells were plated on synthetic dropout nutrient medium (SD/-Ade/-His/-Leu/-Trp) (QDO), and further were incubated on QDO medium containing AbA and X-α-Gal. The interactions between TaPDK and the proteins obtained from positive colonies were further confirmed by co-transformation validation. After plasmids DNA were extracted from blue colonies and sequenced, the sequences results were analyzed by bioinformatic methods. Finally, 24 colonies were obtained, including eight genes, namely non-specific lipid-transfer protein precursor (TanLTP), polyubiquitin (TaPUbi), glyceraldehyde-3-phosphate dehydrogenase, proliferating cell nuclear antigen (TaPCNA), CBS domain containing protein (TaCBS), actin, guanine nucleotide-binding protein beta subunit, chalcone synthase, and three new genes with unknown function. The results of quantitative RT-PCR showed that the expression levels of TanLTP, TaPUbi, and TaPCNA were obviously up-regulated in PHYMS anther, and TaCBS expression was only increased at the tricellular stage in PHYMS anther compared with in fertile lines. Whereas, the expression of TaPDK was obviously down-regulated in PHYMS lines. Collectively, these datas indicated that the majority of candidate proteins might be related to pollen abortion in PHYMS lines, which further suggested that TaPDK plays multiple roles in pollen development, besides participating in regulating pyruvate dehydrogenase complex activity.     

2种玉米青贮饲料青贮过程中主要微生物的变化规律研究

旨在研究玉米青贮过程中各主要微生物的数量变化及变化趋势.做去棒揉丝和带穗切段瓶制青贮,取青贮前玉米青贮原料及在装瓶后第0.5、1、2、3、4、5、6、7、9、11、13、15、20、25、30、35、40、45、50天的青贮样品,用无菌水浸泡,然后用选择性培养基培养其中的主要微生物,包括乳酸菌、酵母菌和霉菌,最后进行计数.结果表明,在玉米青贮密封后60 d内,水分含量变化很小.pH在第2天下降到4以下,然后稳定在3.5左右,带穗切段玉米青贮pH低于去棒揉丝玉米青贮.乳酸菌数量剧增,在第6、7天时达到最高峰,为109数量级,之后数量缓慢下降,于第15～20天时稳定在107数量级,去棒揉丝玉米青贮的乳酸菌数量在第7天达到最高峰,带穗切段玉米青贮则在第6天出现最高峰,其稍高于去棒揉丝玉米青贮.酵母菌在青贮初期数量有些波动,出现最高峰到达107数量级后数量随时间的延长而减少,去棒揉丝玉米青贮时酵母菌数量迅速下降,第40天后没有再检测到酵母菌的存在,而带穗切段玉米在青贮12 h之内数量迅速增加,之后缓慢下降,第50天后没有再检测到酵母菌的存在.霉菌因密封后氧气缺乏而数量很快减少,最晚在第11天左右便检测不到,去棒揉丝玉米青贮中霉菌数量下降较迅速,第4天之后便检测不到霉菌存在,而带穗切段玉米青贮则比较平缓,直到第11天之后便检测不到霉菌存在.综上所述,在玉米青贮过程中主要微生物随时间大致呈现逐步减少的趋势；对玉米秸秆进行纵切揉丝处理有利于玉米青贮饲料的制作.     

Conditional expression of harpinPsscauses yeast cell death that shares features of cell death pathway with harpinPss-mediated plant hypersensitive response (HR)

Conditional expression of harpin_Psscauses yeast cell death that shares features of cell death pathway with harpin_Pss-mediated plant hypersensitive response (HR).Pseudomonas syringae pv.syringae 61 hrp Z gene encodes harpin_Pss, a 34.7 kD extracellular protein that elicits a hypersensitive response (HR) in plants. Conditional expression of either full-length or truncated hrp Z sequences under the GAL1 promoter caused cell death in Saccharomyces cerevisiae Y187. Plating of pYEUT- hrp Z transformants on a medium containing galactose resulted in complete inhibition of colony formation, whereas their growth on a glucose-based medium was unaffected. Western blot analysis confirmed the expression of harpin_Pssin yeast cells transformed with pYEUT- hrp Z and grown in galactose-containing medium. A time-dependent decline in the percentage of trypan blue-excluding cells in cultures of pYEUT- hrp Z transformants was observed when cultured on galactose-containing medium. Similarly, the number of viable cells reduced to about 50% within 6 h. There were similarities in the harpin_Pss-mediated cell death in plants and yeast cell death (YCD). Galactose-induced cell death in pYEUT-hrp Z transformants of S. cerevisiae Y187 was suppressed by a protein kinase inhibitor K252a (10 μ M). The viability of pYEUT- hrp Z transformants was prolonged in the presence of 100 U ml⁻¹catalase suggesting a role for the oxidative burst in YCD that was further supported by the flow cytometric patterns of propidium iodide uptake by yeast cells. Overall, it appears that yeast provides a useful model system to understand the molecular mechanism of harpin_Pss-mediated cell death.     

利用甘蔗糖蜜生产饲料酵母发酵条件的研究

以甘蔗糖蜜为碳源发酵生产高生物量饲料酵母,通过单因素正交试验设计,确定优化的培养条件为:温度28℃,pH5.0,装液量200mL/500mL,摇床转速180r/min,培养时间84h。在优化的培养条件下酵母菌株的总蛋白质含量可达到6.01g/L。     

有机铬对热应激蛋鸡生产性能和血液生化指标的影响

试验选用91周龄新罗曼商品蛋鸡480只,按2×2双因子试验设计随机分为5组,对照组饲喂不添加铬的玉米-豆粕型基础日粮,4个试验组饲喂在基础日粮中分别添加了以吡啶羧酸铬或酵母铬来源的0.2mg/kg或0.4mg/kg的日粮,试验30d,以研究有机铬对热应激蛋鸡生产性能和血液生化指标的影响。试验结果显示:①有机铬能在一定程度上促进热应激蛋鸡的蛋白质利用,提高热应激蛋鸡的生产性能;②铬水平对血清尿素氮有极显著的影响(P<0.01),添加0.4mg/kg铬组均极显著地低于对照组(P<0.01),说明了补铬能够抑制由高温引起的蛋白质分解过程,从而起到缓解高温应激的作用;③吡啶羧酸铬和酵母铬在作用效果上无显著的差异;从经济效益方面考虑,0.4mg/kg铬水平的酵母铬和吡啶羧酸铬经济效益较高。     

Effects of Yeast Culture on Growth Performance,Meat Quality and Intestinal Microflora of Broilers

The experiment was conducted to study the effects of yeast culture on growth performance, meat quality and intestinal microflora of broilers. A total of 432 1-day-old AA commercial broiler chicks with initial body weight of (41.73±0.15)g were randomly divided into 4 groups with 6 replicates per group and 18 broilers per replicate,receiving diets supplemented with 0,0.1%,0.2% and 0.3% yeast culture, respectively. The trial lasted for 42 days which was divided into two periods with 1 to 21 d and 22 to 42 d,respectively.The feed intake,body weight,death and culling rate were recorded in each period. At the end of the experiment,1 chickens in each replicate with the weight close to the average value were slaughtered after fasting for 12 h. The slaughter performance and intestinal microflora counts were determined.The results showed that compared with control group,the supplementation of 0.2% and 0.3% yeast culture significantly increased the body weight at 42 d and the average daily gain (ADG) during the whole trial (P< 0.05).The diet supplemented with 0.2% yeast culture significantly improved the ADG,decreased the F/G of broilers during 22 to 42 d (P< 0.05),and it also significantly increased the average daily feed intake (ADFI) and ADG (P< 0.05),decreased the F/G of the whole period (P< 0.05).Moreover,the supplementation of 0.2% yeast culture significantly decreased the drip loss of meat and the count of Escherichia coli (P< 0.05),and significantly increased the count of Bifidobacterium(P< 0.05). In summary, the supplementation of 0.2% yeast culture could significantly improve the growth performance and the muscle quality of broilers.The intestinal microflora in broiler cecum were effectively optimized after inclusion of yeast culture in the study.