离子交换法精制丙氨酸的研究

合成法生产的丙氨酸,产品中Ｃｌ￣－,含量过高,仅为工业级。我们采用多柱串联的离子交换系统,对工业级丙氨酸进行了脱ＣＬ￣－,脱试验,交换后丙氨酸产品中的Ｃｌ￣－、含量分别小于０．０２％和０．０３％,符合食品添加剂的标准。     

运用ELISA快速检测CPV抗体方法的建立与均衡性研究

用蔗糖密度梯度离心和凝胶层析纯化犬细小病毒 (CPV)作抗原 ,建立了检测CPV抗体的间接酶联免疫吸附试验 (ELISA)法 ,其特异性和稳定性良好 ,结果判定准确明显 ,易于把握。     

玉米-大豆间作和施氮对玉米产量及农艺性状的影响

为研究玉米-大豆间作模式和施氮水平对玉米产量、主要农艺性状及生长动态的影响,进行2个种植模式(玉米单作和玉米-大豆间作)和2个施氮水平(0 kg/hm2,150 kg/hm2)的双因素随机区组试验,以期揭示施氮和间作对玉米产量的影响规律,为提高玉米-大豆间作系统产量提供一定的理论依据。研究结果表明:(1)与不施氮相比,施氮显著增加了春秋两季间作玉米产量,分别达到23.81%和40.99%。施氮处理下的间作玉米地上部生物量较不施氮提高了29.91%,单作模式下显著提高了40.34%,两者差异均达到显著水平。(2)与不施氮相比,施氮150 kg/hm2条件下春玉米单作和间作模式百粒重分别提高了18.92%和19.23%,秋玉米单作和间作模式百粒重分别提高了31.03%和32.75%,差异均达到显著水平。与不施氮相比,施氮150 kg/hm2条件下,单作和间作模式均显著提高秋玉米穗长。与不施氮相比,施氮150 kg/hm2条件下,单作秋玉米的穗粗提高了18.67%,差异显著。(3)施氮和间作均能促进玉米干物质累积、提高株高和叶绿素(SPAD值),且表现为施氮效果高于间作效果。总体来看,种植模式和施氮水平对玉米产量、主要农艺性状和生长动态均有一定影响,且施氮效果优于间作效果。由于土壤具有一定的供氮能力,而间作豆科能为玉米供给一定量的氮素,故对于春玉米而言,施氮效果仅在百粒重中表现,随着土壤原有氮素被玉米吸收利用减少后,供氮能力下降,在秋玉米中施氮效果显著提高。     

Functional Mechanism of Antimicrobial Peptide Bomidin and Its Safety for Macrobrachium rosenbergii

Macrobrachium rosenbergii is an economically important source of crustacean seafood worldwide. Vibrio parahaemolyticus is an important aquatic pathogen that causes epidemics of acute hepatopancreatic necrosis in shrimp populations, which results in significant economic losses to aquaculture farmers. To prevent the antibiotics abuse, which has become a serious threat to human health, novel anti-infective strategies are urgently required to control V. parahaemolyticus. Antimicrobial peptides, which exhibit favourable germicidal activity compared to traditional antibiotics, can be used as a key method to prevent and treat bacterial diseases. Herein, an antimicrobial peptide, bomidin, was expressed through genetic engineering technology. The minimum inhibitory concentration (MIC) of bomidin showed a significant inhibitory effect on V. parahaemolyticus that was equivalent to that of ampicillin. Subsequently, the mechanism of action of recombinant bomidin was explored using PNP and ONPG assays to investigate the effects on membrane permeability. These assays indicated that bomidin penetrated the germ membrane and induced the release of cytoplasmic contents and ultimately interacted with DNA to form a bomidin–DNA complex that inhibits bacterial survival. Transmission electron microscopy and scanning electron microscopy revealed that bomidin could cause damage and dysfunction to the cell wall and membrane. Bomidin was nontoxic to mouse red blood cells within a concentration range that was much larger than the MIC. Toxicity assays revealed that 0.02 mg/mL bomidin was safe for use with juvenile freshwater prawns of M. rosenbergii and significantly inhibited the growth of V. parahaemolyticus in cultured water. These results demonstrated that synthetic peptide bomidin had great antibacterial effect against V. parahaemolyticus and therefore a therapeutic potential in aquaculture.

     

Development and optimization of two-dimensional-electrophoresis protocol ofLeptospirillum ferriphilum

Leptospirillum ferriphilum is important in bioleaching, in which process it is often under heavy stresses of heavy metal ions and high oxidation reduction potential (ORP). Two-dimensional-electrophoresis (2-DE) and comparative proteomic analysis are useful to investigate the responses ofL. ferriphilum to environmental stresses. But, 2-DE analysis forL. ferriphilum is not successful as the samples ofL. ferriphilum contain low protein concentration, complex composition, high salt concentration, and many other interfering components, which make it difficult for 2-DE analysis. In this research, optimizations on the sample preparation and purification methods, sample volume, sample loading methods for isoelectric focusing (IEF), and gel visualization methods were made. More than 629 Coomassie stained spots in single gel were obtained. The image quality and protein concentration in most of the spots met the requirements for both differential spots analysis and mass-spectrum analysis. The 2-DE protocol forL. ferriphilum was successfully developed for the first time.     

浸矿细菌——喜温硫杆菌研究进展

摘要：生物冶金技术具有良好的发展前景。喜温硫杆菌是生物冶金过程中的优势浸矿菌之一,在混合浸矿体系中可协助铁氧化细菌显著提高浸矿效率。本文在分析相关文献的基础上,并结合本实验室的研究工作,从喜温硫杆菌在浸矿过程中的作用机制、抗砷机制、基因组学研究和遗传改造等方面的研究进展进行了综述,并对未来相关研究方向做了展望,旨在为喜温硫杆菌研究提供参考。     

A novel real-time quantitative PCR method using attached universal template probe

A novel real-time quantitative polymerase chain reaction (PCR) method using an attached universal template (UT) probe is described. The UT is an approximately 20 base attachment to the 5′ end of a PCR primer, and it can hybridize with a complementary TaqMan probe. One of the advantages of this method is that different target DNA sequences can be detected employing the same UT probe, which substantially reduces the cost of real-time PCR set-up. In addition, this method could be used for simultaneous detection using a 6-carboxy-fluorescein-labeled UT probe for the target gene and a 5-hexachloro-fluorescein-labeled UT probe for the reference gene in a multiplex reaction. Moreover, the requirement of target DNA length for UT–PCR analysis is relatively flexible, and it could be as short as 56 bp in this report, suggesting the possibility of detecting target DNA from partially degraded samples. The UT–PCR system with degenerate primers could also be designed to screen homologous genes. Taken together, our results suggest that the UT–PCR technique is efficient, reliable, inexpensive and less labor-intensive for quantitative PCR analysis.     

Complete genome of <Emphasis Type="Italic">Leptospirillum ferriphilum</Emphasis> ML-04 provides insight into its physiology and environmental adaptation

Leptospirillum ferriphilum has been identified as the dominant, moderately thermophilic, bioleaching microorganism in bioleaching processes. It is an acidic and chemolithoautrophic bacterium that gains electrons from ferrous iron oxidation for energy production and cell growth. Genetic information about this microorganism has been limited until now, which has hindered its further exploration. In this study, the complete genome of L. ferripilum ML-04 is sequenced and annotated. The bacterium has a single circular chromosome of 2,406,157 bp containing 2,471 coding sequences (CDS), 2 rRNA operons, 48 tRNA genes, a large number of mobile genetic elements and 2 genomic islands. In silico analysis shows L. ferriphilum ML-04 fixes carbon through a reductive citric acid (rTCA) cycle, and obtains nitrogen through ammonium assimilation. The genes related to “cell envelope biogenesis, outer membrane” (6.9%) and “DNA replication, recombination and repair” (5.6%) are abundant, and a large number of genes related to heavy metal detoxification, oxidative and acidic stress defense, and signal transduction pathways were detected. The genomic plasticity, plentiful cell envelope components, inorganic element metabolic abilities and stress response mechanisms found the base for this organism’s survival in the bioleaching niche.     

Instability of the octarepeat region of the human prion protein gene

Prion diseases are a family of unique fatal transmissible neurodegenerative diseases that affect humans and many animals. Sporadic Creutzfeldt-Jakob disease (sCJD) is the most common prion disease in humans, accounting for 85–90% of all human prion cases, and exhibits a high degree of diversity in phenotypes. The etiology of sCJD remains to be elucidated. The human prion protein gene has an octapeptide repeat region (octarepeats) that normally contains 5 repeats of 24–27 bp (1 nonapeptide and 4 octapeptide coding sequences). An increase of the octarepeat numbers to six or more or a decrease of the octarepeat number to three is linked to genetic prion diseases with heterogeneous phenotypes in humans. Here we report that the human octarepeat region is prone to either contraction or expansion when subjected to PCR amplification in vitro using Taq or Pwo polymerase and when replicated in wild type E. coli cells. Octarepeat insertion mutants were even less stable, and the mutation rate for the wild type octarepeats was much higher when replicated in DNA mismatch repair-deficient E.coli cells. All observed octarepeat mutants resulting from DNA replication in E.coli were contained in head-to-head plasmid dimers and DNA mfold analysis (http://mfold.rna.albany.edu/?q=mfold/DNA-Folding-Form) indicates that both DNA strands of the octarepeat region would likely form multiple stable hairpin structures, suggesting that the octarepeat sequence may form stable hairpin structures during DNA replication or repair to cause octarepeat instability. These results provide the first evidence supporting a somatic octarepeat mutation-based model for human sCJD etiology: 1) the instability of the octarepeat region leads to accumulation of somatic octarepeat mutations in brain cells during development and aging, 2) this instability is augmented by compromised DNA mismatch repair in aged cells, and 3) eventually some of the octarepeat mutation-containing brain cells start spontaneous de novo prion formation and replication to initiate sCJD.