Molecular Cloning of a Cellulose Synthase Gene PtoCesA1 from Populus tomentosa and Its Genetic Transformation in Tobacco

A 3 125 bp cellulose synthase gene, PtoCesA1, which has a 98% identity to PtrCesA1 from Populus tremuloides, was cloned from cDNA prepared from secondary xylem of P tomentosa. Four anti-expression vectors with different fragments of PtoCesAl, named as pBIPF, pBICC1, pBIPR and pBIBR, were constructed. Some traits of transformed tobacco of pBICC1, pBIPR and pBIBR differed from wild types, such as small leaves, ＂dwarf＂ phenotype and thinner xylem and fiber cell walls than wild plants consistent with a loss of cellulose. It indicated that the growth of transgenic tobacco was restrained by the expression of anti-PtoCesA1. Transgenic tobacco was obtained and the contents of cellulose and lignin were analyzed as well as the width and length of fiber cells, and xylem thickness for both transgenic and control plants. Transformed tobacco showed a different phenotype from control plants and it implied that PtoCesA1 was essential for the cellulose biosynthesis in poplar stems.     

油茶查尔酮合酶和异构酶基因的cDNA克隆

查尔酮合酶和查尔酮异构酶是类黄酮代谢与色素苷代谢的关键酶.以油茶近成熟种子cDNA文库和EST文库为材料,通过分子克隆方法鉴定了1条查尔酮合酶基因全长cDNA和1条查尔酮异构酶全长cDNA.结果表明:油茶查尔酮合酶基因的cDNA含1479 bp,编码412个氨基酸,为目前最长的查尔酮合酶,与其它物种的查尔酮合酶具有极高的相似性,在进化上高度同源;油茶查尔酮异构酶基因的cDNA含899 bp,编码206个氨基酸,与茶的查尔酮异构酶基因高度同源,但与其它物种的相似性较低.     

生物活性物质角鲨烯的资源及其应用研究进展

通过查阅大量的文献资料，综述了生物活性物质角鲨烯的来源和应用。角鲨烯广泛存在于动植物体内，在动物中以深海鱼类，特别是鲨鱼的肝脏中含量最为丰富，在鲨鱼肝油中的最高含量可达69％；而在植物中则以苋属植物的种子油中含量最高，达5％～8％，在橄榄油中检测到的最高含量为1．16％。角鲨烯有较强的生物活性，被广泛应用在医疗保健品和化妆品等方面。角鲨烯易于氧化，贮存时需要加入适当的抗氧剂。     

Local pulmonary immune responses in domestic cats naturally infected with Cytauxzoon felis

Cytauxzoonosis is a hemoprotozoal disease of cats and wild felids in the South and Southeastern United States caused by Cytauxzoon felis. Although the causative agent has been recognized since the seventies, no study has examined the local immune response in affected organs, such as the lung, and compared them to the lungs of uninfected domestic cats. Previous studies have suggested that the histopathologic findings in the lungs of C. felis-infected cats are caused by the release of pro-inflammatory mediators, such as cytokines and increased production of inducible nitric oxide synthase (iNOS), by the infected macrophages. Our laboratory had previously found an upregulation of the adhesion molecule CD18, which can stimulate the release of these pro-inflammatory mediators. The objective of this study was to characterize local pulmonary immune responses in cats naturally infected with C. felis. Immunohistochemistry was performed to detect tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, iNOS, and major histocompatibility complex (MHC) II in 19 lungs from affected cats that died between 2005 and 2013. Results showed increased expression of all of these molecules when compared to lungs from uninfected, healthy cats. Furthermore, MHC II is expressed in the endothelium of C. felis naturally infected cats. These results support that there is a marked, local, pro-inflammatory immune response that can contribute to the pathogenesis of cytauxzoonosis in the lungs.     

Cross-resistance profile of mesosulfuron-methyl-resistant Italian ryegrass in the southern United States

Diclofop-resistant Lolium species (ryegrass) is a major weed problem in wheat production worldwide. This study was conducted to determine the resistance pattern of diclofop-resistant ryegrass accessions from the southern United States to mesosulfuron-methyl, a recently commercialized herbicide for ryegrass control in wheat; to determine the cross-resistance pattern of a Lolium multiflorum Lam. (Italian ryegrass) accession, 03-1, to acetolactate synthase (ALS) and acetyl-CoA carboxylase (ACCase) inhibitors; and to determine the resistance mechanism of Italian ryegrass to mesosulfuron-methyl. Seventeen ryegrass accessions from Arkansas and Louisiana, including standard resistant and susceptible accessions, were used in this experiment. Fourteen of the 17 accessions were more resistant (four- to > 308-fold) to diclofop than the standard susceptible biotype. One accession, 03-1, was resistant to mesosulfuron-methyl as well as to other ALS inhibitor herbicides such as chlorsulfuron, imazamox and sulfometuron. Accession 03-1, however, did not show multiple resistance to the ACCase inhibitor herbicides diclofop, fluazifop, clethodim, sethoxydim and pinoxaden, nor to glyphosate. The in vivo ALS activity of the 03-1 biotype was less affected by mesosulfuron-methyl than the susceptible biotype. This indicates that the resistance mechanism of Italian ryegrass to mesosulfuron-methyl is partly due to an alteration in the target enzyme, ALS. It is concluded that diclofop-resistant ryegrass in the southern United States can be generally controlled by mesosulfuron-methyl. However, mesosulfuron-methyl must be used with caution because not all ryegrass populations are susceptible to it. There is a need for more thorough profiling of ryegrass resistance to herbicides.     

昆虫几丁质合成关键酶功能及其RNAi技术在害虫防治中的研究进展

几丁质对昆虫的生长发育至关重要,且不存在于植物和哺乳动物中,因此其合成途径的关键酶是较为理想的杀虫剂靶标。近年来,多种害虫的几丁质合成关键酶基因被克隆和鉴定,通过RNAi技术应用于一些重要害虫的防治研究工作中。本文较为系统的总结了近些年害虫几丁质合成关键酶的基因克隆及功能研究进展,重点阐述了几丁质合成酶、海藻糖酶基因基于RNAi技术应用于害虫防治取得的研究进展,分析了RNAi技术通过转基因植物表达dsRNA（RNAi抗虫作物）以及作为核酸农药（dsRNA）直接进行作物喷施的两种应用途径,文章最后还探讨了RNAi在害虫防治中面临的主要瓶颈问题以及主要应对策略。上述较为系统的归纳和总结,目的是为今后基于几丁质合成关键酶的RNAi技术应用于害虫绿色防控研究提供有价值的理论指导。     

多花水仙HMGS基因的克隆

以多花水仙花瓣抑制消减杂交文库获得的羟甲基戊二酰辅酶A合酶（HMGS）基因片段为基础,采用cDNA末端快速扩增（RACE）技术从黄花水仙2号和金盏银台中各克隆一条HMGS基因,两基因均含有一个1413bp的开放阅读框,编码470个氨基酸,但存在2个氨基酸差异．氨基酸序列分析表明：黄花水仙2号与葡萄、大豆、蓖麻和玉米HMGS基因的氨基酸相似系数分别为83％、82％、82％和81％．以水仙Actin基因为内参基因,采用荧光定量PCR方法分析HMGS基因在两品种的表达水平,结果表明：两品种HMGS基因的表达水平差异并不明显．     

栉孔扇贝神经节一氧化氮合酶的组织化学和免疫组化定位

采用组织化学和免疫组化技术对栉孔扇贝(Chlamys farreri)神经节内的一氧化氮合酶(NOS)进行定位研究。组织化学显示,存在NOS的部位如下：脑神经节内纵行的神经纤维和表层的少量小细胞;足神经节表层的大量小细胞,中央大量水平分布的神经纤维;脏神经节中部大量水平分布的神经纤维,前叶内大量小细胞和神经纤维,后叶内少量小细胞和许多环行神经纤维,侧叶内大量似放射状分布的神经纤维;脑足和脑脏神经索内的神经纤维。免疫组化定位表明,神经型一氧化氮合酶(nNOS)和诱导型一氧化氮合酶(iNOS)在整个神经系统内均呈阴性;足神经节和脏神经节内有少量神经细胞呈内皮型一氧化氮合酶(eNOS)强阳性;各神经节和神经索内的部分小细胞和神经纤维呈eNOS弱阳性。栉孔扇贝进化上为较低等的贝类,NOS阳性神经细胞应主要分布于外周器官组织内。神经系统内大量的NOS可在其神经传导和免疫调节等方面发挥重要的作用。     

Role of ILK siRNA on expression of GSK-3β and β-catenin in human tubular epithelial cells stimulated by high glucose

AIM：To investigate the effects of siRNA targeting integrin-linked kinase (ILK) on the expression of glycogen synthase kinase 3β (GSK-3β) and β-catenin during epithelial-mesenchymal transition (EMT) in human kidney proximal tubular epithelial cell line HKC induced by high glucose. METHODS：HKC cells were divided into 4 groups:normal glucose (NG) group, high glucose (HG) group, HG+HK (a vector containing the non-specific siRNA designed as negative control) group and HG＋ILK siRNA group. The inverted fluorescence microscope was used to examine the expression of green fluorescent protein (GFP). The expression of ILK at mRNA and protein levels was detected by RT-PCR and Western blotting. The expression of p-GSK-3β and β-catenin was observed by immunocytochemical staining. The protein expression of total GSK-3β, p-GSK-3β, nuclear β-catenin, total β-catenin, E-cadherin and α-smooth muscle actin (α-SMA) was measured by Western blotting. RESULTS：GFP was observed in HKC cells, indicating that the transfection was successful. Both the protein and mRNA of ILK were down-regulated in HG＋ILK siRNA group compared with HG group and HG+HK group, but still higher than those in NG group. Silencing of ILK down-regulated the expression of p-GSK-3β and nuclear β-catenin. No difference of total GSK-3β or total β-catenin was observed among the 4 groups. CONCLUSION:These data support a functional role of ILK, GSK-3β and β-catenin in tubular EMT induced by high glucose. ILK may promote tubular EMT by regulating the activity of GSK-3β and β-catenin, the downstream effectors of the Wnt/β-catenin pathway.     

Aortic expression of HSP22, TNF-α and eNOS in rats with hyperlipidemia and effects of atorvastatin

AIM:To establish a rat hyperlipidemia model for studying the aortic expression of heat shock protein 22 (HSP22), tumor necrosis factor alpha (TNF-α) and endothelial nitric oxide synthase (eNOS) and the effect of atorvastatin intervention. METHODS:Hyperlipidemia model was established in SD rats. Afterwards, the rats were divided into normal control group, high fat group and high fat+atorvastatin intervention group. The expression of HSP22 and TNF-α in the rat aortas was detected by immunohistochemical assay and the expression of eNOS was assessed by Western blotting. RESULTS:No detectable expression of HSP22 and TNF-α in the normal control group was observed. However, the expression of HSP22 and TNF-α was positive in the high fat group and the atorvastatin intervention group. The mean densities of HSP22 and TNF-α positive particles were significant lower in the atorvastatin intervention group as compared with high fat group (both P＜0.05). The expression of eNOS protein in the high fat group and atorvastatin intervention group was significantly lower than that in normal control group (P<0.01). However, no marked difference of eNOS protein expression between high fat group and atorvastatins intervention group was observed. CONCLUSION: The expression of HSP22 and TNF-α in the rat aortas is increased in the hyperlipidemia rat model. This effect can be restored by atorvastatin treatment. The expression of eNOS in the rat aortas is decreased in the hyperlipidemia rat model, but this tendency could not be attenuated by atorvastatin.