The feasibility of an approximate irregular field dose distribution simulation program applied to a respiratory motion compensation system

PurposeThis study optimized our previously proposed simulation program for the approximate irregular field dose distribution (SPAD) and applied it to a respiratory motion compensation system (RMCS) and respiratory motion simulation system (RMSS). The main purpose was to rapidly analyze the two-dimensional dose distribution and evaluate the compensation effect of the RMCS during radiotherapy.MethodsThis study modified the SPAD to improve the rapid analysis of the dose distribution. In the experimental setup, four different respiratory signal patterns were input to the RMSS for actuation, and an ultrasound image tracking algorithm was used to capture the real-time respiratory displacement, which was input to the RMCS for actuation. A linear accelerator simultaneously irradiated the EBT3 film. The gamma passing rate was used to verify the dose similarity between the EBT3 film and the SPAD, and conformity index (CI) and compensation rate (CR) were used to quantify the compensation effect.ResultsThe Gamma passing rates were 70.48–81.39% (2%/2mm) and 88.23–96.23% (5%/3mm) for various collimator opening patterns. However, the passing rates of the SPAD and EBT3 film ranged from 61.85% to 99.85% at each treatment time point. Under the four different respiratory signal patterns, CR ranged between 21% and 75%. After compensation, the CI for 85%, 90%, and 95% isodose constraints were 0.78, 0.57, and 0.12, respectively.ConclusionsThis study has demonstrated that the dose change during each stage of the treatment process can be analyzed rapidly using the improved SPAD. After compensation, applying the RMCS can reduce the treatment errors caused by respiratory movements.     

基于EMA-qPCR的茄科青枯菌活体检测技术的建立

【目的】利用特异性核酸染料叠氮溴乙锭(Ethidium monoazide bromide, EMA)与实时荧光定量PCR技术相结合, 建立一种能有效区分青枯菌死活细胞的检测方法。【方法】样品DNA制备前经EMA渗透预处理, 再进行实时荧光定量PCR特异扩增菌体DNA。【结果】终浓度为2.0 mg/L的EMA能有效排除1.0×107 CFU/mL灭活青枯菌细胞DNA的扩增, 对活细胞和不可培养状态(Viable but non-culturable, VBNC)活菌的DNA扩增均没有影响。当每个定量PCR反应体系中的活细胞在5.0×100?5.0×104 CFU范围内时, 扩增Ct值与定量PCR反应体系中活细胞CFU对数值呈良好的负相关性(R2=0.992 5)。比较EMA-qPCR法和平板计数法对经过不同温度短期保存的青枯菌检测结果发现, 待检样品可在24 °C与4 °C冷藏条件下短期保存。【结论】本研究建立的EMA-qPCR方法能有效检测青枯菌VBNC细胞和有效区分死活菌, 避免或减少青枯菌PCR检测的假阳性和假阴性。     

油棕（Elaeis guineensis）中果皮发育过程中miRNA的表达动态分析

以CTAB法提取油棕（Elaeis guineensis）中果皮5个不同发育时期（G1～G5）的小RNA。从前期研究获得的油棕小RNA测序数据库中筛选12个候选miRNA,实时荧光定量PCR法（qRT-PCR）检测其在果实发育过程中的表达量变化,并进一步对显著差异表达的miRNA进行靶基因预测。结果表明：中果皮5个不同发育时期小RNA的OD260/OD280比值在1.7～2.0之间;浓度分别是289、364、476、213、390 ng/μL;qRT-PCR检测结果显示,12个候选miRNA在5个发育时期均显著性差异表达,特别是在中果皮发育第4个时期（G4）和第5个时期（G5）表达量极显著增高,其中miR395和miR156在第4个时期表达量最高;miR395和miR528在发育第5时期表达量最高;靶基因预测结果显示差异表达的部分miRNA,其靶基因可能参与了脂肪酸代谢通路,如磷脂酸磷酸脂酶和磷脂酶D。本研究筛选的与脂肪酸代谢相关的miRNA为今后油棕脂肪酸代谢调控通路研究提供了可能的线索。     

Combining ethidium monoazide treatment with real-time PCR selectively quantifies viable Batrachochytrium dendrobatidis cells

Detection of the lethal amphibian fungus Batrachochytrium dendrobatidis relies on PCR-based techniques. Although highly accurate and sensitive, these methods fail to distinguish between viable and dead cells. In this study a novel approach combining the DNA intercalating dye ethidium monoazide (EMA) and real-time PCR is presented that allows quantification of viable B. dendrobatidis cells without the need for culturing. The developed method is able to suppress real-time PCR signals of heat-killed B. dendrobatidis zoospores by 99.9 % and is able to discriminate viable from heat-killed B. dendrobatidis zoospores in mixed samples. Furthermore, the novel approach was applied to assess the antifungal activity of the veterinary antiseptic F10^® Antiseptic Solution. This disinfectant killed B. dendrobatidis zoospores effectively within 1 min at concentrations as low as 1:6400.     

Polymorphism of Fecundity Genes (FecB,FecX, and FecG) in the Indian Bonpala Sheep

The 37-kDa laminin receptor precursor/67-kDa laminin receptor (LRP/LR, also known as ribosomal protein SA, RPSA) has been reported to be involved in cancer development and prion internalization. Previous studies have shown that the LRP/LR is expressed in a wide variety of tissues. In particular, expression of LRP/LR mRNA may be closely related to the degree of PrP^Sc propagation. This study presents a detailed investigation of the LRP/LR mRNA expression levels in eleven normal ovine tissues. Using real-time quantitative PCR, the highest LRP/LR expression was found in neocortex (p < 0.05). Slightly lower levels were found in the heart and obex. Intermediate levels were seen in hippocampus, cerebellum, spleen, thalamus, mesenteric lymph node, and the lowest levels were present in liver, kidney, and lung. In general, the LRP/LR mRNA levels were much higher in neuronal tissues than in peripheral tissues. The observation that differences in LRP/LR mRNA expression levels are consistent with the corresponding variation in PrP^Sc accumulation suggests that the 37-kDa/67-kDa laminin receptor may be involved in the regulation of PrP^Sc propagation.     

实时定量PCR构建法夫酵母内参基因β-actin、 gpd、18S rRNA标准品质粒和标准曲线

为建立检测法夫酵母JMU-MVP14中虾青素合成相关基因在不同生长时期表达水平的实时定量PCR方法,构建法夫酵母JMU-MVP14的管家基因β-actin、gpd、18S rRNA的标准质粒,进行实时定量PCR,制作标准曲线及回归方程.β-actin基因标准曲线相关系数（R2）=0.9956,扩增效率（E） =96.93％;gpd基因标准曲线相关系数（R2） =0.9901,扩增效率（E） =93.78％;18S rRNA基因标准曲线相关系数（R2） =0.9981,扩增效率（E）=98.76％.3个基因片段的熔解曲线均呈单峰;扩增曲线呈典型的S型动力学曲线,指数期和平台期明显,为理想的熔解曲线和扩增曲线.用geNorm软件对三个管家基因的稳定性进行分析,三个基因的稳定性排序为β-actin＞ 18S rRNA＞ gpd,故β-actin和18S rRNA较适合作为研究法夫酵母JMU-MVP14定量实验的内参基因.     

Process analytical technologies to monitor the liquid phase of anaerobic cultures

Recent advances in sensor development and miniaturization offer new possibilities to monitor and control bioprocesses. Specific requirements for anaerobic processes in terms of low costs and high robustness against insoluble fractions and impurities in media led to a decelerated penetration of new technology in this field. Since no regulatory framework demands for process monitoring and documentation like in the pharmaceutical industry, the implementation of new sensors beyond long-established methods is not conducted as intensively.Nevertheless, many attempts have been made in recent years to adopt sensors and show their applicability in anaerobic fermentation processes. New possibilities arise for improved monitoring, control and faster process development through digitalization. This review aims to provide an overview over these recent attempts with the focus on the liquid phase in the upstream part of pure culture bioprocesses for anaerobic bulk compound, food, and beverage production. In particular, methods that monitor the viability, metabolic activity or related parameters are discussed, like electrochemical, impedance and spectroscopy probes, and methods related to fluid flow and gradient formation, like acoustic and mobile sensors.     

Distribution of Karlodinium veneficum in the coastal region of Xiangshan Bay in the East China Sea,as detected by a real-time quantitative PCR assay of ribosomal ITS sequence

Athecate dinoflagellate Karlodinium veneficum is a universal toxic species possessing karlotoxins recognized especially as ichthyotoxic as well as cytotoxic and hemolytic. Blooms of K. veneficum, both single-species or accompanied with other species, occurred more frequently worldwide in recent years, including the coastal region of China. Normally, K. veneficum present in relatively low abundance in phytoplankton communities in estuary regions. Being small and difficult to identify with light microscopy, it has been ignored for a long time till its blooming and toxins being confirmed. How it presents in background level and what is its relationship with critical geological and hydrological environment factors are basically not clear. In this study, the paper reports the application of a real-time quantitative PCR (qPCR) method to investigate the abundance and distribution of K. veneficum in the coastal waters of Xiangshan Bay in the East China Sea (ECS), a typical bay area of harmful algae blooms and heavily affected by anthropogenic activities. The real-time qPCR assay came out being an efficient method at detecting even low cell densities of K. veneficum of different genotypes. A total of 38 field samples of surface (0.5 m) and bottom water (9–100 m in depth) were analyzed and 12 samples were found positive for K. veneficum. At least 3 genotypes of K. veneficum present in this region. Temperatures in sites of K. veneficum positive ranged from 21.7 to 23.4 °C, and salinity levels were between 21.1 and 26.3. The K. veneficum distributed quite extensively in the waters of Xiangshan Bay, cell abundance varied from a low of 4 cells/L to a maximum of 170 cells/L. Most of the samples containing K. veneficum were collected from bottom water in different sites. At three of the 19 sampling sites, K. veneficum was detected in both surface and bottom water samples. Especially at sampling site near Beilun port, where the water is typically muddy with low transparency, relative high cell numbers of K. veneficum were found in both surface and bottom waters. Mixotrophy and vertical migration of K. veneficum could be important eco-physiological factors to consider in terms of understanding these distribution characteristics. The ideal conditions for K. veneficum growth and aggregation in this area still needs further study.