The cell agglutination agent, phytohemagglutinin-L, improves the efficiency of somatic nuclear transfer cloning in cattle (Bos taurus)

One of the several factors that contribute to the low efficiency of mammalian somatic cloning is poor fusion between the small somatic donor cell and the large recipient oocyte. This study was designed to test phytohemagglutinin (PHA) agglutination activity on fusion rate, and subsequent developmental potential of cloned bovine embryos. The toxicity of PHA was established by examining its effects on the development of parthenogenetic bovine oocytes treated with different doses (Experiment 1), and for different durations (Experiment 2). The effective dose and duration of PHA treatment (150 microg/mL, 20 min incubation) was selected and used to compare membrane fusion efficiency and embryo development following somatic cell nuclear transfer (Experiment 3). Cloning with somatic donor fibroblasts versus cumulus cells was also compared, both with and without PHA treatment (150 microg/mL, 20 min). Fusion rate of nuclear donor fibroblasts, after phytohemagglutinin treatment, was increased from 33 to 61% (P < 0.05), and from 59 to 88% (P < 0.05) with cumulus cell nuclear donors. The nuclear transfer (NT) efficiency per oocyte used was improved following PHA treatment, for both fibroblast (13% versus 22%) as well as cumulus cells (17% versus 34%; P < 0.05). The cloned embryos, both with and without PHA treatment, were subjected to vitrification and embryo transfer testing, and resulted in similar survival (approximately 90% hatching) and pregnancy rates (17-25%). Three calves were born following vitrification and embryo transfer of these embryos; two from the PHA-treated group, and one from non-PHA control group. We concluded that PHA treatment significantly improved the fusion efficiency of somatic NT in cattle, and therefore, increased the development of cloned blastocysts. Furthermore, within a determined range of dose and duration, PHA had no detrimental effect on embryo survival post-vitrification, nor on pregnancy or calving rates following embryo transfer.     

Analysis of the downstream region of nodD3 P1 promoter by deletion and complementation tests in Sinorhizobium meliloti

Rhizobia specifically interacts with the host, leguminous plants, leading to the formation of nitrogen-fixing root nodules. The Rhizobium genes essential for nodule formation are called nodulation genes (nod or nol). The expression of nod genes requires the presence of host signals, generally flavonoids, and the product of regulatory nodD gene, NodD[1,2]. The expression of nod genes results in the synthesis of Nod factors, which serve as the signal molecules to elicit root cor-tical cells di…     

三七总RNA提取方法的对比研究

比较利用改进的异硫氰酸胍一步法、异硫氰酸胍高盐法、CTAB法和Thomas’RNA提取法等4种方法提取三七根茎2个部位总RNA的可行性。结果表明,改进的异硫氰酸胍一步法和异硫氰酸胍高盐法能有效地抑制酚类物质、多糖及皂苷等次级代谢产物对总RNA的影响,可从三七根茎中获得质量高、完整性好的总RNA。RT—PCR分析显示提取的总RNA具有反转录活性。这2种方法具有快速、简单、有效的特点。     

红豆杉生物工程的研究进展

自从紫杉醇(taxol,又称paclitaxel,结构见图1)被批准作为植源性抗癌药应用以来,由于其天然资源的限制,使得近几年来对寻找及扩大紫杉醇药源途径的研究取得了极大的进展。这些途径大致包括:(1)筛选高产量红豆杉(Taxus)栽培品种,(2)化学合成,(3)生物技术,(4)微生物生产。在这些研究领域特别是生物技术研究领域中,由于目前对紫杉醇的生物合成及关键酶研究所取得的进展,已使人们相信通过基因工程手段作为最佳生产紫杉醇药源途径之一,在不久的将来将会变为现实。本文报道的就是有关紫杉醇在生物工程方面的研究进展概况。1　研究简史红豆杉体外培…     

Characterization and function analysis of a cold-induced AmCIP gene encoding a dehydrin-like protein in Ammopiptanthus mongolicus.

A cDNA clone was isolated after difference screening from cotyledons of two-week cold-treated Ammopiptanthus mongolicus. The full-length cDNA sequence [designated as AmCIP (A. mongolicus cold-induced protein) gene] was 806 bp long and contained a 465 bp open reading frame (ORF) encoding a 16.6 kD protein of 154 amino acids. Bioinformatic analyses indicated that CIP belongs to dehydrin family with the features of high hydrophilicity, a helix K-segment, a long Gly-rich region and a phosphorylatable tract of Ser as well as deficiency in Cys and Trp. The expression of CIP gene increased after two weeks of cold treatment and more expression was detected in radicle than in cotyledon. And PCR amplification of the AmCIP gene from genome of A. mongolicus revealed this gene has no intron. Function prediction suggested this protein seems to protect the stabilization of membrane structure and prevent macromolecular coagulation or sequestrate calcium ions by association or disassociation with membrane under low temperature conditions.     

Facilitation promotes invasions in plant‐associated microbial communities

While several studies have established a positive correlation between community diversity and invasion resistance, it is less clear how species interactions within resident communities shape this process. Here, we experimentally tested how antagonistic and facilitative pairwise interactions within resident model microbial communities predict invasion by the plant–pathogenic bacterium Ralstonia solanacearum. We found that facilitative resident community interactions promoted and antagonistic interactions suppressed invasions both in the lab and in the tomato plant rhizosphere. Crucially, pairwise interactions reliably explained observed invasion outcomes also in multispecies communities, and mechanistically, this was linked to direct inhibition of the invader by antagonistic communities (antibiosis), and to a lesser degree by resource competition between members of the resident community and the invader. Together, our findings suggest that the type and strength of pairwise interactions can reliably predict the outcome of invasions in more complex multispecies communities.     

LC-MS/MS determination of helicid in human plasma and its application in pharmacokinetic studies

Helicid is a traditional Chinese medicine used to treat headache and insomnia with definite effects. To facilitate pharmacokinetic studies of helicid in man, a sensitive and specific LC-MS/MS method for the quantitative detection of helicid in human plasma was developed and validated. The method involved the addition of bergeninum as the internal standard (IS), protein precipitation, HPLC separation, and quantification by MS/MS system using negative electrospray ionization in the multiple reaction monitoring mode (MRM). The precursor→product ion transitions were monitored at m/z 282.8→120.9 for helicid and m/z 326.9→192.2 for the IS, respectively. The lower limit of quantification (LLOQ) was 0.2 μg/L. The calibration curves for helicid was linear over a concentration range of 0.2-20 μg/L. The intra- and inter-batch analyses of QC samples at 0.4, 2, 20 μg/L indicated good precision (%R.S.D. between 2.69 and 5.47%) and accuracy (between 96.15 and 105.05%). The helicid was stable in human plasma stored at room temperature for at least 24h, 4°C for at least 24h, -20°C for at least 1 month, and for routine three freeze-thaw cycles. This accurate and specific assay provides a useful method for evaluating the pharmacokinetic profile of helicid in humans.     

Integrated soil and plant phosphorus management for crop and environment in China. A review

In semi-arid grassland ecosystems, soil biogeochemical processes are controlled by seasonal and inter-annual rainfall variation and temperature, which may override the long-term impact of grazers on N availability and N dynamics. In a three-year (2004?C2006) case study of an Inner Mongolian grassland, we analysed time-integrated (ion-exchange resins) and instantaneous (soil mineral N extractions) inorganic N availability at three sites of varying grazing intensities and combined these data with information on soil water content (SWC), aboveground net primary productivity (ANPP) and plant N uptake. Additionally, the effects of rainfall and grazing on N-form availability (NO ₃ ^? -N, NH ₄ ⁺ -N) were considered. Grazing had less impact on N availability compared to seasonal and annual rainfall distribution. One of the three study years (2004) showed a grazing effect with higher resin-N availability at the ungrazed site compared to the heavily grazed site. Inorganic N availability was low in the driest year (2005) and highest in a year of average rainfall amount and favourable distribution (2004). In general, we found a positive relationship between inorganic N availability and both plant productivity and plant N uptake. Rainfall also controlled the plant available NO ₃ ^? -N and NH ₄ ⁺ -N pools; NH ₄ ⁺ -N dominated the available inorganic N-form in times of low SWC, while the available NO ₃ ^? -N increased with SWC. We observed N availability and plant productivity in a temporal synchronized pattern. Increased rainfall variability and land-use practices affecting SWC will likely alter N availability dynamics (and the relation of N-forms) and, therefore, important processes of semi-arid natural grassland carbon and N cycling.     

Large-scale identification of proteins in human salivary proteome by liquid chromatography/mass spectrometry and two-dimensional gel electrophoresis-mass spectrometry

Human saliva contains a large number of proteins and peptides (salivary proteome) that help maintain homeostasis in the oral cavity. Global analysis of human salivary proteome is important for understanding oral health and disease pathogenesis. In this study, large-scale identification of salivary proteins was demonstrated by using shotgun proteomics and two-dimensinal gel electrophoresis-mass spectrometry (2-DE-MS). For the shotgun approach, whole saliva proteins were prefractionated according to molecular weight. The smallest fraction, presumably containing salivary peptides, was directly separated by capillary liquid chromatography (LC). However, the large protein fractions were digested into peptides for subsequent LC separation. Separated peptides were analyzed by on-line electrospray tandem mass spectrometry (MS/MS) using a quadrupole-time of flight mass spectrometer, and the obtained spectra were automatically processed to search human protein sequence database for protein identification. Additionally, 2-DE was used to map out the proteins in whole saliva. Protein spots 105 in number were excised and in-gel digested; and the resulting peptide fragments were measured by matrix-assisted laser desorption/ionization-mass spectrometry and sequenced by LC-MS/MS for protein identification. In total, we cataloged 309 proteins from human whole saliva by using these two proteomic approaches.