玉米地方种质沃日黄与其选系重组自交后代的SSR分子标记分析

为探讨玉米地方种质与其选系重组的育种效果,利用SSR标记,分析了玉米地方种质沃日黄与其选系重组自交后代的遗传关系.结果表明,31对引物均检测到了重组系与沃30不同的基因型;供试材料间的遗传相似系数变幅为0.47～0.84,平均遗传相似系数为0.61,平均值较大,但变幅也较大;聚类分析可将沃30及8个重组系划分为三类,重组系1、2、3、 5与沃30聚为一类,重组系4、8和重组系6、7分别聚为一类.以上结果表明,不仅部分重组系与沃30存在一定的遗传差异,而且重组系彼此间也存在较大的遗传差异,地方种质与其选系重组具有明显的育种效果.此外,从计算每条染色体上相同基因型百分比的平均值发现,染色体1上相同基因型3对引物的平均值仅为16.67%,远远低于基因组上相同基因型的比例52.40%,这是否揭示沃30与沃日黄的基因重组可能较多发生在染色体1上,值得进一步研究.     

重组猪干扰素α的纯化工艺研究

[目的]研究重组猪干扰素α的纯化工艺,为进一步研究该蛋白的分子结构及rPoIFN-α标准品的制备奠定基础。[方法]将含重组菌E.coliBL21诱导表达后得到的粗制rPoIFN-α,分别用2种方案进行纯化。方案1,依次用GST亲和层析法、DEAE阴离子交换法及分子筛法层析三步法进行纯化;方案2,依次用GST亲和层析和分子筛法层析两步法进行纯化。纯化结果用SDS-PAGE及Western-blot进行纯度检测及鉴定。[结果]诱导表达后的表达产物经SDS-PAGE检测,发现在45Kd分子量处有目的条带,Western-blot检测有特异性条带。通过两步法纯化后的rPoIFN-α纯度达96%,蛋白得率为42.8%,三步纯化纯度为98.8%。[结论]两步法得到的蛋白纯度非常接近三步法,但与三步法相比工艺更为简单方便。     

添加前缀对重组蛋白生物学特性影响的预测分析

利用分子生物学软件DNASTAR对DAB389-αMSH和含有亲和标记、Xa factor识别序列、连接肽的DAB389（Gly4Ser）2-αMSH的表位、亲水性、等电点等生物学特性进行了预测分析。结果显示：DAB389（Gly4Ser）2-αMSH的表位为0.8,DAB389-αMSH为2.1;亲水性为2.7,高于DAB389-αMSH的1.96;两者的等电点均为5.9。His.Tag、Xa factor识别序列使重组蛋白的亲水性增强,表位降低;重组蛋白氨基端前70个氨基酸中不存在信号肽;DAB389（Gly4Ser）2-αMSH的表达量低于DAB389-αMSH2%左右,前缀对蛋白表达量有一定的影响。     

蔗糖浓度对渗透休克法提取细胞周质重组蛋白的影响

[目的]研究不同蔗糖浓度的高渗液对渗透休克法(Osmotic Shock)提取细胞周质蛋白的影响。[方法]将已经构建好的含Fbp A基因的表达质粒转化到EC感受态细胞中,分别在20%、30%、40%和50%的蔗糖高渗液条件下用Osmotic Shock纯化蛋白,SDS-PAGE电泳检测蛋白。[结果]30%和40%的蔗糖高渗液能得到浓度更高的重组蛋白,而用20%和50%的蔗糖高渗液提取蛋白产量较低。[结论]30%和40%的蔗糖高渗液能提高Osmotic Shock法提取细胞周质重组蛋白产量。     

Re-emerging of porcine respiratory and reproductive syndrome virus (lineage 3) and increased pathogenicity after genomic recombination with vaccine variant

Porcine reproductive and respiratory syndrome virus (PRRSV) was first reported in China since late 1995 and several variants were further reported in subsequence years, causing huge economic losses to the Chinese swine industry. To date, three major lineages (lineage 3, 5.1 and 8.7) of Type 2 PRRSV were reported in China based on our global genotyping. The present study provides the epidemiology of the PRRSV in South China based on the isolates collected during 2009–2012, indicating three lineages (lineage 3, 5.1 and 8.7) of Type 2 PRRSV were still circulating in this area. Our phylogenetic reconstruction indicated that lineage 3 re-emerged in 2010 formed a huge cluster with closely related to the 2004 isolates from Hong Kong. Furthermore, the inter-lineage genomic recombination between MLV vaccine strain (lineage 5) and a recently re-emerged lineage 3 virus (QYYZ) has also been found in a farm practicing MLV vaccination. Our in vivo experiment comparing the pathogenicity and clinical presentations among currently isolated viruses indicated that pigs infected with recombinant lineage 3 virus (GM2) showed persistent higher fever compared to pigs infected by its wild counterpart (QYYZ). This study enhanced our understanding on potential importance of the recombination of PRRSV along with their evolution.     

植物减数分裂重组的影响因素·机制及在育种中的应用

总结了影响减数分裂重组频率的诸多因素,探讨了提高减数分裂重组频率对植物远缘杂交中不良基因与优异基因的不利连锁打破的可能,并就当前的问题和今后的发展前景进行了讨论。     

玉米籽粒生理成熟后自然脱水速率QTL的初步定位

以吉846(脱水快,1.18%d-1)和掖3189(脱水慢,0.39%d-1)为亲本衍生出的232个重组自交系(F7)为作图群体,构建了具有101个SSR标记位点的玉米遗传连锁图谱,覆盖玉米基因组1941.7cM,标记间平均距离为19.22cM。通过1年2点试验(双城和哈尔滨,2007年)评价了232个重组自交系籽粒生理成熟后的自然脱水速率。采用WinQTL2.5对该性状数量性状位点(QTL)进行了初步定位和遗传效应分析,结果共检测出9个显著影响玉米籽粒生理成熟后自然脱水速率的QTL,分别位于第2、第3、第4、第5和第6染色体上,加性增效作用均来源于亲本吉846。其中在第2和第6染色体上的2个QTL(qKdr-2-1和qKdr-6-1)在2个环境下均稳定表达,分别位于SSR标记bnlg198～umc1516之间和phi126～phi077之间,其累积表型贡献率为15.49%。具有较快脱水速率的等位基因均来自吉846。所检测到的QTL将在分子辅助选育具有较快脱水速率的材料中具有较大的应用潜力。     

酶解条件对几种植物性饲料降解效果的影响

用自制的2种多酶组分、高酶活力的复合酶制剂,在水浴40℃条件下,设置了不同pH缓冲液(pH3.5,4.8和6.0),不同酶解时间(0.5,6,12,24h),酶解大麦、玉米、豆粕3种植物性饲料的试验研究。结果表明:①添加酶制剂的酶解还原糖得率均高于不加酶处理,不同酶制剂和不同饲料间存在较大差异;②酶解反应12h时,3种饲料的还原糖得率均达到最高;③大麦、玉米在缓冲液pH4.8条件下酶解效果最好,而豆粕在pH4.8和6.0时没有明显差异。     

Construction of the recombinant baculovirus AcMNPV with cathepsin B-like proteinase and its insecticidal activity against Helicoverpa armigera

The Helicoverpa armigera cathepsin B-like proteinase (HCB) has been shown to have a wide spectrum of substrates. It has been involved in the degradation of yolk protein during embryonic development and also in the decomposition of the adult fat body. To study the possibility of using HCB to improve the insecticidal activity of bioinsecticides, it was inserted into the pFASTBACDUAL-green fluorescent protein (GFP) donor plasmid under the strong polyhedrin promoter, and the polyhedrin gene was retained behind the HCB gene so that the virus could orally infect the host and survive in the natural environment. After the recombinant plasmid transfected the Sf21 cells, the recombinant baculovirus Autographa californica multiple nucleocapsid nucleopolyhedrovirus (AcMNPV)-GFP-HCB-Polh⁺ was produced. A 37-kDa recombinant procathepsin B was expressed in AcMNPV-GFP-HCB-Polh⁺-infected Sf21 cells and processed into 29/25-kDa mature forms. Gelatin zymography revealed that the proteolytic activities of the recombinant HCB and other proteases were activated or enhanced by HCB in AcMNPV-GFP-HCB-Polh⁺-infected larvae. Green fluorescence was observed earlier and was more intense in AcMNPV-GFP-HCB-Polh⁺-infected larvae than in HCB-free AcMNPV-GFP-Polh⁺-infected ones that were infected at the same time; this indicated that the AcMNPV-GFP-HCB-Polh⁺ virus spreaded faster in larvae than the AcMNPV-GFP-Polh⁺ virus. A bioassay revealed that infection with the AcMNPV-GFP-HCB-Polh⁺ virus shortened the median survival time of H. armigera larvae by 12 h as compared with those infected with the AcMNPV-GFP-Polh⁺ or the wild-type AcMNPV virus. These results suggest that HCB may possibly play a role in the recombinant virus in improving the rate of killing larvae.     

黑木耳重组食品的研究

为充分利用木耳碎片及其根部，同时研究多种多糖肢体复配在特定条件下凝胶的特性及效果，使木耳碎片或其根部重新组合，形成不同形状、强度产品，改变其原来口味，易于进食，并更加富于营养，节省了大量木耳，减少资源浪费。