影响棉铃虫增殖马尾松毛虫质型多角体病毒的条件因子的初步探讨

棉铃虫为马尾松毛虫多角体病毒 (DPCPV)理想的替代寄主。试验结果表明 ,不同接毒方法、接种浓度、接种温度及接种虫龄对棉铃虫增殖病毒有影响 ,以病毒涂在人工饲料表面的接毒方法、2 0～ 2 3℃温度及 3龄虫为最佳组合条件。由于利用棉铃虫生产出的病毒 (Ha DPCPV)容易诱发棉铃虫本身的多角体病毒 (NPV) ,尤其对那些带有NPV源的虫种 ,Ha DPCPV不宜作毒种 ,因此为慎重起见 ,最好用马尾松毛虫复制出的病毒作为生产毒种。     

伤寒沙门菌非编码RNA617的分子鉴定及其对生物膜形成的影响研究

本研究旨在探讨伤寒沙门菌(Salmonella enterica serovar Typhi, S. Typhi)中非编码RNA617(non-coding RNA617,ncRNA617)的分子特性,并研究其对生物膜形成的影响及作用机制。采用Northern blot方法检测ncRNA617的表达,通过cDNA 5’末端快速扩增技术(5’-rapid amplification of cDNA end,5’RACE)和逆转录-聚合酶链式反应(reverse transcriotion-polymerase chain reaction,3’RT-PCR)实验分析ncRNA617可能的转录起始位点和终止位点;构建ncRNA617缺陷菌株、回补菌株和过表达菌株等相关菌株,通过生物膜形成实验,观察ncRNA617对伤寒沙门菌生物膜形成的影响,并用实时荧光定量聚合酶链式反应(quantitative real-time polymerase chain reaction,qPCR)分析生物膜形成相关基因表达水平的变化,综合运用生物信息学方法预测ncRNA617和差异基因的结合区域,初步分析ncRNA617发挥调控作用的机制。结果显示,伤寒沙门菌确有ncRNA617的表达,长度约300 nt,其转录起始位点位于mig-14终止密码子下游967 nt处,终止位点位于t2681起始密码子上游 2 378～2 560 nt处。与野生对照菌株相比,ncRNA617缺陷菌株生物膜形成能力增强(P<0.05),回补菌株的生物膜形成能力恢复至野生菌株水平,过表达菌株的生物膜形成能力有所下降(P<0.05)。qPCR结果表明,ncRNA617可负向调控多个生物膜形成相关基因的转录表达水平(P<0.05)。经生物信息学方法预测发现,ncRNA617与差异基因有不同的结合区域。本研究结果提示,ncRNA617在伤寒沙门菌中存在,其长度约270～452 nt。ncRNA617可能通过靶向结合生物膜形成相关基因下调基因表达,从而负向调控伤寒沙门菌生物膜的生成。     

The NSR1 gene encodes a protein that specifically binds nuclear localization sequences and has two RNA recognition motifs

We previously identified a protein (p67) in the yeast, Saccharomyces cerevisiae, that specifically recognizes nuclear localization sequences. We report here the partial purification of p67, and the isolation, sequencing, and disruption of the gene (NSR1) encoding this protein. p67 was purified using an affinity column conjugated with a peptide containing the histone H2B nuclear localization sequence from yeast. Using antibodies against p67 we have cloned the gene for this protein. The protein encoded by the NSR1 gene recognizes the wild-type H2B nuclear localization sequence, but does not recognize a mutant H2B sequence that is incompetent for nuclear localization in vivo. Interestingly, the NSR1 protein has two RNA recognition motifs, as well as an acidic NH2 terminus containing a series of serine clusters, and a basic COOH terminus containing arg-gly repeats. We have confirmed the nuclear localization of p67 by immunofluorescence and found that a restricted portion of the nucleus is highlighted. We have also shown that NSR1 (p67) is required for normal cell growth.     

Functional analysis of slow myosin heavy chain 1 and myomesin-3 in sarcomere organization in zebrafish embryonic slow muscles

Myofibrillogenesis, the process of sarcomere formation, requires close interactions of sarcomeric proteins and various components of sarcomere structures. The myosin thick filaments and M-lines are two key components of the sarcomere. It has been suggested that myomesin proteins of M-lines interact with myosin and titin proteins and keep the thick and titin filaments in order. However, the function of myomesin in myofibrillogenesis and sarcomere organization remained largely enigmatic. No knockout or knockdown animal models have been reported to elucidate the role of myomesin in sarcomere organization in vivo. In this study, by using the gene-specific knockdown approach in zebrafish embryos, we carried out a loss-of-function analysis of myomesin-3 and slow myosin heavy chain 1 (smyhc1) expressed specifically in slow muscles. We demonstrated that knockdown of smyhc1 abolished the sarcomeric localization of myomesin-3 in slow muscles. In contrast, loss of myomesin-3 had no effect on the sarcomeric organization of thick and thin filaments as well as M- and Z-line structures. Together, these studies indicate that myosin thick filaments are required for M-line organization and M-line localization of myomesin-3. In contrast, myomesin-3 is dispensable for sarcomere organization in slow muscles.     

Expression of Tumor-Associated Macrophage in Progression of Human Glioma

The aim of this study is to investigate the expression of tumor-associated macrophages (TAMs) M1, M2 phenotypic in human glioma tissues, and to explore the clinical significance and prognostic value of TAMs in glioma patients. A total of 50 glioma samples were obtained from patients diagnosed in our hospital from 2007 to 2010. Clinical follow-up was conducted via return visits and telephone interviews after discharge. Progression free survival (PFS) was calculated based on tumor progression by MRI and CT examination from the primary operation. Overall survival (OS) time was calculated from the initial surgical operation date to end date of follow-up or death. Kaplan–Meier methodology was used to evaluate the survival of patients and log-rank test for comparing differences between groups. The expression levels of CD16 and CD206 were investigated in the 4 μm serial paraffin sections by immunohistochemistry. M1-type macrophages filtrated in all the grades of glioma samples, and the lower expression level was associated with high grade glioma. A negative correlation was found between WHO pathological grades and the expression of M1-type macrophages by Spearman correlation analysis. M2-type macrophages filtrated in all the grades of glioma samples with the higher expression level associated with high grade glioma. A positive correlation was found between WHO pathological grades and the expression of M2-type macrophages by Spearman correlation analysis. The PFS and OS among patients with high levels of M1-type macrophages (CD16+++) were significantly higher than those with less expression. The PFS and OS among patients with high levels of M2-type macrophages (CD206+++) were significantly lower than those with low expression. M1-type macrophages may inhibit the tumor growth and improve the therapeutic outcome of glioma patients. M2 ratios are associated with tumor proliferation and poor prognosis. TAMs phenotypes of glioma samples are the potential biomarkers in assessing the degree of malignancy, tumor invasion, and patient prognosis in clinic.     

iTRAQ-based proteomics analysis of autophagy-mediated immune responses against the vascular fungal pathogen Verticillium dahliae in Arabidopsis

The mechanisms underlying the functional link between autophagy and plant innate immunity remain largely unknown. In this study, we investigated the autophagy-mediated plant defense responses against Verticillium dahliae (V. dahliae) infection by comparative proteomics and cellular analyses. An assessment of the autophagy activity and disease development showed that autophagic processes were tightly related to the tolerance of Arabidopsis plant to Verticillium wilt. An isobaric tags for relative and absolute quantification (iTRAQ)-based proteomics analysis was performed, and we identified a total of 780 differentially accumulated proteins (DAPs) between wild-type and mutant atg10-1 Arabidopsis plants upon V. dahliae infection, of which, 193 ATG8-family-interacting proteins were identified in silico and their associations with autophagy were verified for several selected proteins. Three important aspects of autophagy-mediated defense against V. dahliae infection were revealed: 1) autophagy is required for the activation of upstream defense responses; 2) autophagy-mediated mitochondrial degradation (mitophagy) occurs and is an important player in the defense process; and 3) autophagy promotes the transdifferentiation of perivascular cells and the formation of xylem hyperplasia, which are crucial for protection against this vascular disease. Together, our results provide several novel insights for understanding the functional association between autophagy and plant immune responses.     

种子物理休眠研究进展

种子物理休眠是由种皮不透水层引起的一种休眠类型,是植物在长期系统发育进程中获得的一种适应环境变化的特性。该文简述了种子物理休眠的定义与概念;从不透水层、种皮的特殊水孔器结构以及胚的形态特异性等方面,综述了物理休眠种子的形态特征、物理休眠与综合休眠的解除方法以及物理休眠的可能解除机制;利用Angiosperm Phylogeny Group Ⅲ(APG Ⅲ)系统分析了种子物理休眠的植物在系统发育中的位置;最后提出了今后种子物理休眠有待研究的主要问题。     

A strong root-specific expression system for stable transgene expression in bread wheat

Key message

A strong, stable and root-specific expression system was developed from a rice root-specific GLYCINE - RICH PROTEIN 7 promoter for use as an enabling technology for genetic manipulation of wheat root traits.

Abstract

Root systems play an important role in wheat productivity. Genetic manipulation of wheat root traits often requires a root-specific or root-predominant expression system as an essential enabling technology. In this study, we investigated promoters from rice root-specific or root-predominant expressed genes for development of a root expression system in bread wheat. Transient expression analysis using a GREEN FLUORESCENT PROTEIN (GFP) reporter gene driven by rice promoters identified six promoters that were strongly expressed in wheat roots. Extensive organ specificity analysis of three rice promoters in transgenic wheat revealed that the promoter of rice GLYCINE-RICH PROTEIN 7 (OsGRP7) gene conferred a root-specific expression pattern in wheat. Strong GFP fluorescence in the seminal and branch roots of wheat expressing GFP reporter driven by the OsGRP7 promoter was detected in epidermal, cortical and endodermal cells in mature parts of the root. The GFP reporter driven by the promoter of rice METALLOTHIONEIN-LIKE PROTEIN 1 (OsMTL1) gene was mainly expressed in the roots with essentially no expression in the leaf, stem or seed. However, it was also expressed in floral organs including glume, lemma, palea and awn. In contrast, strong expression of rice RCg2 promoter-driven GFP was found in many tissues. The GFP expression driven by these three rice promoters was stable in transgenic wheat plants through three generations (T1–T3) examined. These data suggest that the OsGRP7 promoter can provide a strong, stable and root-specific expression system for use as an enabling technology for genetic manipulation of wheat root traits.