A tailing genome walking method suitable for genomes with high local GC content

The tailing genome walking strategies are simple and efficient. However, they sometimes can be restricted due to the low stringency of homo-oligomeric primers. Here we modified their conventional tailing step by adding polythymidine and polyguanine to the target single-stranded DNA (ssDNA). The tailed ssDNA was then amplified exponentially with a specific primer in the known region and a primer comprising 5′ polycytosine and 3′ polyadenosine. The successful application of this novel method for identifying integration sites mediated by φC31 integrase in goat genome indicates that the method is more suitable for genomes with high complexity and local GC content.     

Structural analysis of Canavalia maritima and Canavalia gladiata lectins complexed with different dimannosides: new insights into the understanding of the structure-biological activity relationship in legume lectins

Plant lectins, especially those purified from species of the Leguminosae family, represent the best studied group of carbohydrate-binding proteins. The legume lectins from Diocleinae subtribe are highly similar proteins that present significant differences in the potency/efficacy of their biological activities. The structural studies of the interactions between lectins and sugars may clarify the origin of the distinct biological activities observed in this high similar class of proteins. In this way, this work presents a crystallographic study of the ConM and CGL (agglutinins from Canavalia maritima and Canavalia gladiata, respectively) in the following complexes: ConM/CGL:Man(alpha1-2)Man(alpha1-O)Me, ConM/CGL:Man(alpha1-3)Man(alpha1-O)Me and ConM/CGL:Man(alpha1-4)Man(alpha1-O)Me, which crystallized in different conditions and space group from the native proteins. The structures were solved by molecular replacement, presenting satisfactory values for R(factor) and R(free). Comparisons between ConM, CGL and ConA (Canavalia ensiformis lectin) binding mode with the dimannosides in subject, presented different interactions patterns, which may account for a structural explanation of the distincts biological properties observed in the lectins of Diocleinae subtribe.     

Automatic segmentation of surface EMG images: Improving the estimation of neuromuscular activity

Surface electromyograms (EMGs) recorded with a couple of electrodes are meant to comprise representative information of the whole muscle activation. Nonetheless, regional variations in neuromuscular activity seem to occur in numerous conditions, from standing to passive muscle stretching. In this study, we show how local activation of skeletal muscles can be automatically tracked from EMGs acquired with a bi-dimensional grid of surface electrodes (a grid of 8 rows and 15 columns was used). Grayscale images were created from simulated and experimental EMGs, filtered and segmented into clusters of activity with the watershed algorithm. The number of electrodes on each cluster and the mean level of neuromuscular activity were used to assess the accuracy of the segmentation of simulated signals. Regardless of the noise level, thickness of fat tissue and acquisition configuration (monopolar or single differential), the segmentation accuracy was above 60%. Accuracy values peaked close to 95% when pixels with intensity below ～70% of maximal EMG amplitude in each segmented cluster were excluded. When simulating opposite variations in the activity of two adjacent muscles, watershed segmentation produced clusters of activity consistently centered on each simulated portion of active muscle and with mean amplitude close to the simulated value. Finally, the segmentation algorithm was used to track spatial variations in the activity, within and between medial and lateral gastrocnemius muscles, during isometric plantar flexion contraction and in quiet standing position. In both cases, the regionalization of neuromuscular activity occurred and was consistently identified with the segmentation method.     

Purification and molecular cloning of a new galactose-specific lectin from <Emphasis Type="Italic">Bauhinia variegata</Emphasis> seeds

A new galactose-specific lectin was purified from seeds of a Caesalpinoideae plant, Bauhinia variegata, by affinity chromatography on lactose-agarose. Protein extracts haemagglutinated rabbit and human erythrocytes (native and treated with proteolytic enzymes), showing preference for rabbit blood treated with papain and trypsin. Among various carbohydrates tested, the lectin was best inhibited by D-galactose and its derivatives, especially lactose. SDS-PAGE showed that the lectin, named BVL, has a pattern similar to other lectins isolated from the same genus, Bauhinia purpurea agglutinin (BPA). The molecular mass of BVL subunit is 32 871 Da, determined by MALDI-TOF spectrometry. DNA extracted from B. variegata young leaves and primers designed according to the B. purpurea lectin were used to generate specific fragments which were cloned and sequenced, revealing two distinct isoforms. The bvl gene sequence comprised an open reading frame of 876 base pairs which encodes a protein of 291 amino acids. The protein carried a putative signal peptide. The mature protein was predicted to have 263 amino acid residues and 28 963 Da in size.     

小麦-中间偃麦草双体异附加系的选育和鉴定

在小麦-中间偃麦草59个杂交后代种质系中,筛选出6个小麦-中间偃麦草双体异附加系(line0605,line0607,line0609,line0610,line0611,line0625),并对其进行了形态学、白粉病抗性、细胞学和RAPD鉴定。形态学结果表明:6个双体异附加系农艺性状较好地结合了双亲的优良特点;细胞学结果表明:6个双体异附加系具有高度的细胞学稳定性,花粉母细胞减数分裂中期Ⅰ(PMCMI)的染色体构型为2n=22Ⅱ;RAPD分析表明:在供试的209个随机引物中有5个引物分别能在6个异附加系中稳定地扩增出不同的特异带型,可以作为各个异附加系所附加染色体的特异分子标记;白粉病抗性鉴定结果表明:line0605表现免疫,line0610和line0625表现高抗,line0607表现中抗,line0609和line0611表现中感。     

影响山羊体外受精的因素

以屠宰山羊卵母细胞为材料研究了公羊个体、附睾不同部位精子、成熟培养和受精时卵丘存在与否、卵丘扩展程度及卵龄对山羊体外受精的影响。结果表明 :1)不同公羊精液在受精、卵裂和桑椹 /囊胚率上都有显著差异 ;2 )附睾尾精子和鲜精的受精、卵裂和桑椹 /囊胚率无显著差异 ,但显著高于附睾体和附睾头精子 ;3)成熟培养 2 4和 2 7h卵母细胞的的桑椹胚 /囊胚率显著高于培养 2 1和 30h卵母细胞 ;4 )卵丘扩展 3和 4级卵母细胞受精和桑椹胚 /囊胚率显著高于扩展 0和 1级卵母细胞 ;5 )成熟培养前机械去卵丘严重影响卵母细胞体外受精和桑椹胚 /囊胚率 ;6 )受精前完全去掉卵丘显著影响桑椹胚 /囊胚率     

Genetic Variation of the VP1 Gene of the Virulent Duck Hepatitis A Virus Type 1(DHAV-1) Isolates in Shandong Province of China

To investigate the relationship of the variation of virulence and the external capsid proteins of the pandemic duck hepatitis A virus type 1(DHAV-1) isolates,the virulence,cross neutralization assays and the complete sequence of the virion protein 1(VP1) gene of nine virulent DHAV-1 strains,which were isolated from infected ducklings with clinical symptoms in Shandong province of China in 2007-2008,were tested.The fifth generation duck embryo allantoic liquids of the 9 isolates were tested on 12-day-old duck embryos and on 7-day-old ducklings for the median embryonal lethal doses(ELD 50 s) and the median lethal doses(LD 50 s),respectively.The results showed that the ELD 50 s of embryonic duck eggs of the 9 DHAV-1 isolates were between 1.9 × 10 6 /mL to 1.44 × 10 7 /mL,while the LD 50 s were 2.39 × 10 5 /mL to 6.15 × 10 6 /mL.Cross-neutralization tests revealed that the 9 DHAV-1 isolates were completely neutralized by the standard serum and the hyperimmune sera against the 9 DHAV-1 isolates,respectively.Compared with other virulent,moderate virulent,attenuated vaccine and mild strains,the VP1 genes of the 9 strains shared 89.8%-99.7% similarity at the nucleotide level and 92.4%-99.6% at amino acid level with other DHAV-1 strains.There were three hypervariable regions at the C-terminus(aa 158-160,180-193 and 205-219) and other variable points in VP1 protein,but which didn’t cause virulence of DHAV-1 change.     

Variations in the Spatial Distribution of the Amplitude of Surface Electromyograms Are Unlikely Explained by Changes in the Length of Medial Gastrocnemius Fibres with Knee Joint Angle

This study investigates whether knee position affects the amplitude distribution of surface electromyogram (EMG) in the medial gastrocnemius (MG) muscle. Of further concern is understanding whether knee-induced changes in EMG amplitude distribution are associated with regional changes in MG fibre length. Fifteen surface EMGs were acquired proximo-distally from the MG muscle while 22 (13 male) healthy participants (age range: 23–47 years) exerted isometric plantar flexion at 60% of their maximal effort, with knee fully extended and at 90 degrees flexion. The number of channels providing EMGs with greatest amplitude, their relative proximo-distal position and the EMG amplitude averaged over channels were considered to characterise changes in myoelectric activity with knee position. From ultrasound images, collected at rest, fibre length, pennation angle and fat thickness were computed for MG proximo-distal regions. Surface EMGs detected with knee flexed were on average five times smaller than those collected during knee extended. However, during knee flexed, relatively larger EMGs were detected by a dramatically greater number of channels, centred at the MG more proximal regions. Variation in knee position at rest did not affect the proximo-distal values obtained for MG fibre length, pennation angle and fat thickness. Our main findings revealed that, with knee flexion: i) there is a redistribution of activity within the whole MG muscle; ii) EMGs detected locally unlikely suffice to characterise the changes in the neural drive to MG during isometric contractions at knee fully extended and 90 degrees flexed positions; iii) sources other than fibre length may substantially contribute to determining the net, MG activation.     

基于耕地生态足迹的重庆市耕地生态承载力供需平衡研究

以耕地生态承载力供需关系为切入点,应用耕地生态足迹模型和GIS方法,在修正了Wackernagel提出的耕地产量因子的基础上,构建耕地生态承载力供需平衡指数,以2009年为例,对重庆市的耕地生态承载力供需平衡状况进行了系统的评价.结果表明:(1)重庆市耕地生态承载力供需空间分布表现出较为明显的不平衡性,总体上从东到西呈现出从富裕到严重超载的空间分布态势,且耕地生态超载区与主城区分布具有较好的吻合性;(2)就县(区)数量而言,重庆市耕地生态赤字区最大,其次是耕地生态盈余区和耕地生态平衡区;就人口数量而言,同样是耕地生态赤字区最大,其次是耕地生态平衡区和耕地生态盈余区;就土地面积而言,耕地生态盈余区最大,其次是耕地生态平衡区,耕地生态赤字区面积最小.重庆市约3/5的人口分布在约4/5表现为耕地生态平衡或盈余的土地面积上,而约2/5的人口集中分布在约1/5表现为耕地生态赤字的土地面积上.可为重庆市的区域耕地生态安全预警、土地利用总体规划以及区域可持续发展提供科学依据和决策支持.