贵州省猪繁殖与呼吸综合征病毒分子流行病学调查

为了解2007年4月至2008年7月贵州省猪繁殖与呼吸综合征病毒(PRRSV)分子流行病学特征,在此期间基于PRRSV的Nsp2基因在全省33个发病猪场展开调查。结果表明,有3种不同Nsp2基因缺失类型毒株在贵州省流行。参考PRRSV经典毒株相应序列,根据Nsp2基因缺失的不同情况将PRRSV贵州省流行毒株分为3种类型,类型Ⅰ(3/33)缺失177个碱基,类型Ⅱ(29/33)缺失90个碱基,类型Ⅲ(1/33)缺失93个碱基。类型Ⅱ毒株与近年来国内广泛流行的高致病性PRRSV缺失情况一致,类型Ⅰ和Ⅲ是2种新发现的缺失型毒株,但其系统发生地位与高致病性PRRSV接近。     

2013年中国典型对虾养殖区白斑综合征病毒流行株高变异区序列的分析比较

为了解中国不同地区白斑综合征病毒的流行变异情况,本研究取用2013年3—12月从7个省市发病地区采集到的64份WSSV阳性样本,以特异的引物扩增目的片段,通过测序分析不同样本的缺失及变异差异。结果显示,在开放阅读框ORF14/15的扩增中,分别有6530 bp、6533 bp和5138 bp的片段缺失,而在ORF23/24扩增中有12070bp大片段的缺失,不同地区样本中未能成功扩增ORF75,而ORF94的重复单元数目分别为0、3、4、12不等,ORF125的重复单元数目为0、7。SNP分析表明,含有3个重复单元的ORF94在48位的碱基为T、T、T,重复单元数为4的在48位的碱基为T、T、T、T,重复单元数为12的在48位的碱基分别为T及11个A。而ORF125所有重复单元数为7的情况在8、18、25、66、69位置的碱基均为G、G、G、G、A,在9、50、53、61位的碱基也普遍出现了变异。结果表明,WSSV在中国不同地区存在一定程度的变异,其在序列中的缺失、重复单元数目以及SNP的差异较为明显。     

猪种布氏杆菌WboA基因缺失株对绵羊的免疫剂量及免疫程序研究

为优化猪种布氏杆菌WboA基因缺失株(B.suisΔWboA)对绵羊的免疫条件,本研究采用1岁左右的成年雌性绵羊对B.suisΔWboA的免疫剂量及免程序进行了比较研究。实验分5个组进行,其中A、B、C 3个试验组分别以2倍剂量重复接种、4倍剂量重复接种和单剂量1次接种B.suisΔWboA,D组单剂量1次接种猪种布氏杆菌S2疫苗株(B.suis S2),E组为空白对照组。各组羊首免后7 d、21 d和35 d分别采血,测定血清抗体水平;在首免后35 d,分别采用布氏杆菌强毒菌M28株(B.melitensis M28),经腹股沟皮下注射攻毒。攻毒后28 d,分别取试验羊的脾脏分离攻毒菌株。所有试验羊,在实施攻毒前,其精神、食欲均正常。血清抗体测定结果表明,在二免7 d、21 d后,A组和B组试验羊的抗体水平明显高于C组,而且均超过D组试验羊的抗体水平。攻毒后的细菌分离结果表明,攻毒后28 d,A组和B组试验羊的脾脏细菌分离数量明显低于C组试验羊,并且均低于D组试验羊的细菌分离水平。实验结果表明,B.suisΔWboA的免疫剂量由单倍改为2倍或4倍,免疫程序由单剂量1次改为2倍或4倍剂量2次,可以明显提高免疫效果,并达到与亲本疫苗菌株B.suis S2的免疫水平。     

利用基因工程改造瘤胃微生物调控瘤胃发酵研究进展

DNA分子的人工合成是基因工程技术的基础,改变反刍动物瘤胃微生物DNA的结构,能够使得动物表达出不同的蛋白质,反刍动物的生长模式也因此而发生不可预知的改变。正在应用的或处于研究阶段的构建瘤胃基因工程菌的方式主要有基因缺失技术、基因复制性重组技术和启动子的应用技术。其中基因缺失技术还包括转座子插入法和自杀性质粒的构建。目前影响基因工程技术成功率的最主要障碍就在于这种技术的低效率和不可预知性,一旦清除了这个障碍,基因工程技术将会在瘤胃微生物的改造上有更加广阔的发展空间。     

Effect of high-molecular-weight glutenin subunit deletion on soft wheat quality properties and sugar-snap cookie quality estimated through near-isogenic lines

High-molecular-weight glutenin subunits (HMW-GSs) play a critical role in determining the viscoelastic properties of wheat dough. The HMW-GSs are encoded by Glu-A1, Glu-B1, and Glu-D1 loci on the long arms of chromosomes 1A, 1B, and 1D, respectively. In the present study, four near-isogenic lines with different HMW-GS deletions and compositions at the Glu-A1 and Glu-D1 loci in Yangmai 18 background were used for quality analysis. Deletion in Glu-D1 showed much weaker gluten quality and dough strength than null Glu-A1 genotype and wild genotype (WT), based on the measurements of sodium dodecyl sulfate (SDS)-sedimentation, lactic acid solvent retention capacity (SRC), gluten index, development time, stability time, and alveograph P and L values. The deletion of Glu-D1 did not significantly affect grain hardness, grain protein content, water SRC, sodium carbonate SRC, and sucrose SRC. Double null genotype in Glu-A1 and Glu-D1 and single null genotype in Glu-D1 showed significantly higher cookie diameter, crispness, and lower cookie height compared with single null genotype in Glu-A1 and WT. These indicate that the null Glu-D1 genotype is useful for improvement of biscuit quality, and use of this germplasm would be a viable strategy to develop new wheat varieties for biscuit processing.     

β-catenin基因26 bp插入/缺失对陕北白绒山羊生长性状的遗传效应分析

旨在研究β-连环蛋白(CTNNB1)基因上的多态性位点,并与陕北白绒山羊生长性状进行关联分析,寻找与生长性状相关的遗传标记,为山羊高生产力的标记辅助选择提供科学依据。以陕北白绒山羊(n=848)为研究对象,PCR大样本扩增检测β-catenin基因的插入/缺失(InDel)突变,并评估该位点与生长性状的相关性。结果发现,在848只陕北白绒山羊中β-catenin基因的第10内含子上存在一个26bp InDel的位点;该群体中共出现3种基因型,分别为II、ID和DD。关联性分析结果表明,该突变位点与陕北白绒山羊胸深显著相关(P<0.05),其中杂合型ID为优势基因型。因此,β-catenin基因可作为陕北白绒山羊生长性状选育的候选基因,为陕北白绒山羊选育工作提供理论依据。     

In vitro and in vivo efficacy of a live attenuated Salmonella Typhimurium vaccine at preventing intestinal colonization in chicks

Vaccination of chicks with Salmonella (S .) Typhimurium aroA deletion mutants has previously been shown to inhibit intestinal colonization of wild‐type S.  Typhimurium strains. In Australia, Bioproperties VaxSafe? STM1 strain is the only licensed and commercially available S . Typhimurium vaccine. This vaccine is a live attenuated aroA deletion mutant. Currently, it is recommended that the first dose of the STM1 vaccine is administered through coarse spray. It is unclear whether this mode of administration effectively permits intestinal colonization. Furthermore, it is not known whether the STM1 strain prevents or inhibits Salmonella colonization of chicks following this first dose. This study investigated both in vitro and in vivo colonization parameters. Invasiveness was assessed using an in vitro invasion assay into sections of ileum and caecum collected from day‐old chicks. The S.  Typhimurium definitive types (DT) 9 and 44 exhibited the greatest invasion into both intestinal segments. STM1 was invasive but was significantly less so than both isolates of S.  Typhimurium. In dual and triple infections, no competitive microbial interactions between STM1 and wild‐type Salmonella were observed. In vivo colonization inhibition was also tested. Vaccinated and nonvaccinated day‐old chicks were challenged with S.  Typhimurium DT9. Both STM1 and S.  Typhimurium DT9 were found in spleen, liver, ileum, caecum and caecal contents from day 2 postinfection. No significant exclusion effect was observed in vaccinated and challenged chicks.     

Establishing an essential amino acid profile for maintenance in poultry using deletion method

This study aimed to estimate the essential amino acid profile and the ideal ratio for the maintenance of poultry by deletion method. A nitrogen balance (NB) trial was conducted using 198 adult roosters, housed individually in metabolic cages. The treatments were 33 purified diets being 11 diets with an amino acid mixture providing high protein intake of 500 mg N/BW_kg^0.75 per day, 11 diets providing medium protein intake of 250 mg N/BW_kg^0.75 per day (in each diet, one amino acid tested was diluted 50%) and 11 diets providing low protein intake of 125 mg N/BW_kg^0.75 per day (made by omitting the amino acid tested). Each treatment had six replicates. After 48 h of fasting receiving water plus sucrose, the roosters were fed 40 g of the diets by tube once a day for 3 days. The excreta were collected within 72 h after the first feeding. The diets and excreta were analysed for nitrogen content. For each amino acid studied, a linear regression was fitted by NB and amino acid intake (AAI). The maintenance requirements were estimated as the AAI to maintain the NB equal to zero. The daily amino acid requirements for maintenance were estimated to be Lys 11, Met 29, Thr 23, Trp 5, Arg 50, Val 29, His 6, Gly 54, Phe 49, Leu 78 and Ile 21 mg/BW_kg^0.75 per day. Therefore, the amino acid ratio for maintenance was concluded to be Lys 100, Met 276, Thr 220, Trp 48, Arg 467, Val 275, His 60, Gly 511, Phe 467, Leu 735 and Ile 198% independent of the scale. The essential amino acid profile and the ideal ratio for the maintenance of poultry estimated in this study contributed to improve the factorial model for estimating essential amino acid requirements for poultry.     

布鲁氏菌BPE159基因缺失株的构建及BPE159蛋白对细胞自噬因子表达的影响

【目的】构建布鲁氏菌BPE159基因缺失株,研究缺失株体外生长变化特征及其在宿主细胞中的存活能力,探究布鲁氏菌感染期间分泌蛋白BPE159对自噬因子表达的影响。【方法】同源重组方法构建布鲁氏菌BPE159基因重组质粒,电转化布鲁氏菌S2308感受态细胞构建BPE159基因缺失株S2308ΔBPE159。PCR扩增BPE159基因,连接转化构建pBBR1MCS-4-BPE159载体,提取质粒进行电转化,构建BPE159基因回补株S2308ΔBPE159-C。琼脂糖凝胶电泳检测缺失株和回补株遗传稳定性。构建布鲁氏菌感染小鼠巨噬细胞RAW264.7模型,实时荧光定量PCR检测布鲁氏菌侵染后自噬细胞因子ATG5、Beclin1、LC3a和LC3b基因表达水平。以S2308、S2308ΔBPE159和S2308ΔBPE159-C株侵染小鼠巨噬细胞,收集细胞总RNA,实时荧光定量PCR检测BPE159基因缺失对布鲁氏菌侵染后自噬细胞因子表达水平的影响。在相同起始浓度下培养S2308、S2308ΔBPE159及S2308ΔBPE159-C株,观察细菌生长变化趋势;评价S2308ΔBPE159株在不同...