Modifying Myxococcus xanthus protoporphyrinogen oxidase to plant codon usage and high level of oxyfluorfen resistance in transgenic rice

Protoporphyrinogen oxidase (Protox) of Myxococcus xanthus (Mx Protox) is a 49-kDa membrane protein that catalyzes conversion of protoporphyrinogen IX (Protogen IX) into protoporphyrin IX (Proto IX). Upon heterologous expression in transgenic rice plants, Mx Protox is dually targeted into plastids and mitochondria, increasing resistance against the herbicidal Protox inhibitor oxyfluorfen. Here, we describe the chemical synthesis of the Mx Protox gene by assembling several small synthetic DNA fragments derived by ligation-PCR. Codon usage in the resulting 1416-bp gene was modified to correspond to that of the Arabidopsis Protox gene, a change that resulted in a decrease in G+C content from 71 to 49%. The modified Mx Protox gene was used to generate transgenic rice plants via Agrobacterium-mediated transformation. Integration, expression, and inheritance of the transgenes were demonstrated by Southern, Northern, and Western blot analyses. In plants transformed with the modified, low G+C-content Mx Protox gene, levels of Protox expression and enzyme activity were low compared to the levels observed for plants transformed with the native Mx Protox gene. Nonetheless, like the native gene, the modified gene conferred a high level of resistance to the herbicide oxyfluorfen in a seedling growth test.     

香菇多糖对肉鸡生长性能的影响研究

香菇多糖(LNT)和其他多糖一样是极性大分子,其特定结构与生物活性关系密切。尽管有关LNT的成分和结构报道甚多,但已明确结构与免疫活性关系的只有β-葡聚糖物质[1-2]。LNT在动物中的研究证实其具有免疫调节、抗肿瘤、抗感染等作用,因而将LNT作为安全、高效的新型饲料添加剂的研究对畜禽生产具有重要的理论和实际意义。1材料与方法1·1试验材料和试验动物试验材料:香菇多糖,购自上海康周真菌多糖有限公司,香菇多糖含量40%。试验动物:1日龄罗斯308(ROSS 308)肉鸡400只。1.2试验设计和日粮组成1.2·1试验设计选用400只1日龄的ROSS 308…     

添加酒糟和玉米粉对花生壳微贮效果的影响

通过测定单独的花生壳原样及花生壳中添加10%的酒糟或玉米粉经过微贮后其有机物(粗蛋白、粗纤维、粗脂肪、无氮浸出物)及营养价值的变化情况,研究花生壳的微贮效果。试验结果表明:花生壳经过微贮后营养价值有所提高,原料花生壳中CP含量为9.85%,而微贮后纯花生壳中CP含量为10.99%,添加酒糟和玉米粉微贮后的花生壳中CP含量分别为12.13%、13.25%,微贮后单独的花生壳、添加酒糟和玉米粉的花生壳中CP含量和原料花生壳相比分别上升了11.57%、23.15%、34.52%,添加酒糟和玉米粉的花生壳中CP与原样CP含量相比差异显著;原料花生壳中CF含量为70.82%,而微贮后纯花生壳中CF含量为69.44%,添加酒糟和玉米粉微贮后的花生壳中CF含量分别为68.56%、67.41%,微贮后纯花生壳、添加酒糟和玉米粉的花生壳中CF含量和原料花生壳相比分别下降了1.95%、3.19%、4.82%,添加酒糟和玉米粉的花生壳CF与原样CF含量相比差异显著;在花生壳中添加玉米粉效果比添加酒糟效果好。     

扶正解毒散超微粉对人工感染传染性法氏囊病预防试验

以扶正解毒散超微粉1%、0.5%、0.25%浓度拌料给药,通过死亡率、血清中抗体滴度及增重等指标评价预防效果。结果表明:扶正解毒散超微粉高、中剂量组对抗体、脾脏代偿性增重以及平均增重效果均较突出。临床推荐剂量为0.5%浓度拌料,自由采食饮水,连用5 d。     

Enhancement of protective immune responses by oral vaccination with Saccharomyces cerevisiae expressing recombinant Actinobacillus pleuropneumoniae ApxIA or ApxIIA in mice

We previously induced protective immune response by oral immunization with yeast expressing the ApxIIA antigen. The ApxI antigen is also an important factor in the protection against Actinobacillus pleuropneumoniae serotype 5 infection; therefore, the protective immunity in mice following oral immunization with Saccharomyces cerevisiae expressing either ApxIA (group C) or ApxIIA (group D) alone or both (group E) was compared with that in two control groups (group A and B). The immunogenicity of the rApxIA antigen derived from the yeast was confirmed by a high survival rate and an ApxIA-specific IgG antibody response (p < 0.01). The highest systemic (IgG) and local (IgA) humoral immune responses to ApxIA and ApxIIA were detected in group E after the third immunization (p < 0.05). The levels of IL-1β and IL-6 after challenge with an A. pleuropneumoniae field isolate did not change significantly in the vaccinated groups. The level of TNF-α increased in a time-dependent manner in group E but was not significantly different after the challenge. After the challenge, the mice in group E had a significantly lower infectious burden and a higher level of protection than the mice in the other groups (p < 0.05). The survival rate in each group was closely correlated to the immune response and histopathological observations in the lung following the challenge. These results suggested that immunity to the ApxIA antigen is required for optimal protection.     

Isolation and Identification of Enterococcus faecalis from Fur-bearing Animals

In order to obtain Enterococcus faecalis from fur animals and evaluate its prebiotic properties,in this study,Enterococcus faecalis was isolated from the feces of healthy adult fur-bearing animals (mink,fox,raccoon dog),identified by morphological observation,biochemical test and 16S rRNA sequence analysis.The growth curve,acid production capacity and antibiotic sensitivity of the Enterococcus faecalis isolates were measured to evaluate their probiotic properties.Some strains were selected to determine their tolerance to temperature,artificial gastric juice and artificial bile salt.The results showed that five strains were Gram-positive,and their biochemical characteristics were basically consistent with the standard strains of Enterococcus faecalis,and they were identified as Enterococcus faecalis by 16S rRNA sequence analysis.The five strains all entered the logarithmic phase at 2 h after culture,and entered the stable phase at 8-10 h,and had weak acid production capacity.The resistance rate of the isolates to tetracycline and levofloxacin was 100%,followed by penicillin (80%),erythromycin (80%),gentamicin (80%) and chloramphenicol (40%).All the isolates were sensitive to ampicillin and vancomycin.Enterococcus faecalis from mink,fox and raccoon dog had strong tolerance to temperature below 60 ℃,artificial gastric juice with pH>3.0 and 0.3%-0.5% concentration of bile salt,but poor tolerance to temperature above 70 ℃,and artificial gastric juice with pH<3.0.In conclusion,five strains of Enterococcus faecalis from fur animals (mink,fox,raccoon dog) were obtained in this study.The isolated strains propagated rapidly,which were suitable for colonization and played a prebiotic role in fur animals' intestines,and had good prebiotic characteristics and stress resistance.They could be used as candidate strains for animal microbiological agents for further study.     

湖南两江峡谷森林公园夏季鸟类资源调查

2007年7月4～11日,对湖南两江峡谷森林公园夏季鸟类资源进行了调查。通过样线调查法并结合鸟网捕捉,共记录到68种鸟类,隶属于14目31科,占湖南省400种鸟类的17.00%。其中53种属东洋界种类、3种属古北界种类、12种属广布种;留鸟60种、夏候鸟8种。8种鸟类属国家Ⅱ级保护动物,占鸟类物种数的11.76%。根据调查结果与分析,提出了鸟类资源保护与利用的合理性建议。     

品种与季节和日龄对仔猪血清中IgG含量的影响

为了探索品种、季节和日龄对仔猪血清中IgG含量的影响,将280头试验猪按不同品种/类群(久仰香猪、剑白香猪和长白猪)、不同季节和不同日龄(0、4、7、14、21、28日龄)采样、然后用双抗体夹心ELISA法(DAS-ELISA)测定其血清中IgG含量。结果表明:剑白香猪IgG含量比久仰香猪和长白猪高,差异显著(P<0.05);冬季仔猪IgG含量比其他季节高,差异极显著(P<0.01),春季与夏、秋季差异显著(P<0.05);仔猪IgG含量随日龄而增加,但从21日龄开始下降。     

6株猪O型口蹄疫病毒VP1基因的克隆与序列分析

根据口蹄疫病毒（FMDV）VP1基因的序列，设计并合成了2对用于扩增VP1基因的引物。从组织中提取总RNA，首先用P1、P2引物对6株猪。型口蹄疫病毒进行RT—PCR扩增，获得1000bp的片段；再用P3、P4引物进行巢式PCR扩增，结果获得850bp的片段。将850bp的片段克隆到pMD18—-T载体中，通过PCR鉴定，将阳性重组质粒进行测序并分析。结果发现6株FMDV的核苷酸同源性为80．2％～99．4％，其推导的氨基酸序列同源性为86．9％～99．5％；构建遗传发生树，发现6株FMDV属于两个不同的基因型，其中的Shunde00、Sihui01、Shenzhen99、Fushan01株属一个基因型（与Hongkong93、广东86分离株属同一基因型）；Guangzhou99、Shenzhen00株属另一个基因型（与UKG-12—2001株、JPN2000株属同一基因型）。通过对口蹄疫病毒VP1基因的测序与分析，了解其变异情况，为科学地防控FMD提供分子水平的依据。     

牛分枝杆菌特异性PCR检测方法的建立及初步应用

根据已发表的牛分枝杆菌的pncA的基因序列，设计和合成了一对可扩增294bp目的片段的引物，建立了特异性检测牛分枝杆菌的PCR方法。对牛分枝杆菌国际参考株和国内分离株成功扩增出294bp的特异性基因片段；对人结核分枝杆菌、副结核分枝杆菌、鸟胞内分枝杆菌和草分枝杆菌DNA的PCR扩增结果均为阴性。本PCR方法检测的敏感度可达到50pg。对10份牛分枝杆菌培养阳性和10份阴性样品的DNA分别进行了PCR检测，结果10份阳性样品中有9份样品为PCR扩增阳性，阳性符合率为90％（9／10）；而10份阴性样品则PCR扩增全部为阴性，阴性符合率为100％（10／10）。本方法可做为牛分枝杆菌的快速检测和流行病学调查的工具。