scholarly journals Real-time detection of PRT1-mediated ubiquitination via fluorescently labeled substrate probes

2016 ◽  
Author(s):  
Augustin C. Mot ◽  
Erik Prell ◽  
Maria Klecker ◽  
Christin Naumann ◽  
Frederik Faden ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Real Time ◽  
Ad Hoc ◽  
E3 Ligase ◽  
List Type ◽  
Reaction Optimization ◽  
Protein Functions ◽  
Bona Fide ◽  
Turn Over ◽  
Short Time

SUMMARYThe N-end rule pathway has emerged as a major system for regulating protein functions by controlling their turn-over in medical, animal and plant sciences as well as agriculture. Although novel functions and enzymes of the pathway were discovered, ubiquitination mechanism and substrate specificity of N-end rule pathway E3 Ubiquitin ligases remained elusive. Taking the first discovered bona fide plant N-end rule E3 ligase PROTEOLYSIS1 (PRT1) as a model, we use a novel tool to molecularly characterize polyubiquitination live, in real-time.We gained mechanistic insights in PRT1 substrate preference and activation by monitoring live ubiquitination by using a fluorescent chemical probe coupled to artificial substrate reporters. Ubiquitination was measured by rapid in-gel fluorescence scanning as well as in real time by fluorescence polarization.Enzymatic activity, substrate specificity, mechanisms and reaction optimization of PRT1-mediated ubiquitination were investigated ad hoc in short time and with significantly reduced reagent consumption.We demonstrated for the first time that PRT1 is indeed an E3 ligase, which was hypothesized for over two decades. These results demonstrate that PRT1 has the potential to be involved in polyubiquitination of various substrates and therefore pave the way to understanding recently discovered phenotypes of prt1 mutants.

scholarly journals Quickly and simply detection for coronaviruses including SARS-CoV-2 on the mobile Real-Time PCR without treating RNA in advance

2020 ◽  
Author(s):  
Masaaki Muraoka ◽  
Yukiko Tanoi ◽  
Tetsutaro Tada ◽  
Mikio Mizukoshi ◽  
Osamu Kawaguchi
Keyword(s):  
Real Time ◽  
Correlation Coefficient ◽  
Real Time Pcr ◽  
Positive Control ◽  
List Type ◽  
Rt Pcr ◽  
Human Coronavirus ◽  
One Step ◽  
Short Time ◽  
Human Coronavirus 229E

ABSTRACTSARS-CoV-2 was reported to the WHO as an outbreak in Wuhan City, China on end of 2019, afterwards pandemic on the worldwide in 2020. The SARS-CoV-2 virus is less deadly, but far more transmissible. Therefore, it needs to detect and monitor quickly and simply on site to prevent SARS-CoV-2.If detecting coronaviruses including SARS-CoV-2, the real-time RT-PCR method is sensitive and specific for the unique target, however, it must take long time and labour that RNA is treated in advance, transcribed and amplified. Therefore, referenced previously report, in this study, we modified various methods to prove hypotheses the followed.Firstly, we hypothesized that real-time RT-PCR could be finished in very short time by the mobile real-time PCR device and one-step RT-PCR reagent. Secondly, we hypothesized that it was possible to perform RT-PCR utilizing the reagent as the above without RNA treatment in advance so called “direct”.Firstly, it was able to detect the positive control RNA of SARS-CoV-2 for less than 13.5 minutes by primer-probe referring to the CDC. Moreover, each detection value varied in accordance with each concentration (This correlation coefficient R2 > 0.95). Secondary, it was possible to detect human coronavirus 229E with direct RT-PCR. Furthermore, each detection value varied in accordance with each titer (TCID50 / mL) of human coronavirus 229E (This correlation coefficient R2 > 0.95).Considering the above, causing by utilizing the mobile real-time PCR device and the one-step real-time PCR reagent simultaneously following as: 1) It was possible to detect SARS-CoV-2 in very short time as compared to conventional method; 2) It was possible to detect human coronavirus quickly and simply with “direct”. For these reasons, we hypothesized that it is possible to detect SARS-CoV-2 quickly and simply by utilizing methods the above without treating RNA in advance. This hypothesis is our next try.STRENGTHS AND LIMITATIONS OF THIS STUDY*This study developed it possible to detect the positive control RNA of SARS-CoV-2 more quickly than previously, however couldn’t try to detect the genetic RNA.*This study proved clearly that the human coronavirus instead of SARS-CoV-2 could be detected simply without treating RNA in advance by the same method above.*This study couldn’t try to utilize the human specimens because of our institution limited.*This study could utilize the device and the reagents commercial and not especial.

Improved Macro-clusters generation using Top-k shared Micro-clusters in Data Streams

2017 ◽  
Vol 7 (10) ◽  
pp. 52
Author(s):  
LAKSHMI PRANEETHA
Keyword(s):  
Real Time ◽  
Data Streams ◽  
Bloom Filter ◽  
Scientific Applications ◽  
Pruning Algorithm ◽  
Density Data ◽  
Data Points ◽  
Short Time ◽  
Information Streams

Now-a-days data streams or information streams are gigantic and quick changing. The usage of information streams can fluctuate from basic logical, scientific applications to vital business and money related ones. The useful information is abstracted from the stream and represented in the form of micro-clusters in the online phase. In offline phase micro-clusters are merged to form the macro clusters. DBSTREAM technique captures the density between micro-clusters by means of a shared density graph in the online phase. The density data in this graph is then used in reclustering for improving the formation of clusters but DBSTREAM takes more time in handling the corrupted data points In this paper an early pruning algorithm is used before pre-processing of information and a bloom filter is used for recognizing the corrupted information. Our experiments on real time datasets shows that using this approach improves the efficiency of macro-clusters by 90% and increases the generation of more number of micro-clusters within in a short time.

Avenues to Precision Oncology

2020 ◽  
pp. 36-45
Author(s):  
Alexander Meisel
Keyword(s):  
Precision Medicine ◽  
Trial Design ◽  
Ad Hoc ◽  
Molecular Targets ◽  
Clinical Trial Design ◽  
Predictive Biomarkers ◽  
Precision Oncology ◽  
Companion Diagnostics ◽  
Bona Fide ◽  
The One

Until recently, the clinical management of cancer heavily relied on anatomical and histopathological criteria, with ad hoc guidelines directing the therapeutic choices in specific indications. In the last years, the development and therapeutic implementation of novel anticancer therapies significantly improved the clinical outcome of cancer patients. Nonetheless, such cutting-edge approaches revealed the limitation of the one-size-fits-all paradigm. The newly discovered molecular targets can be exploited either as bona fide targets for subsequent drug development, or as tools to precision medicine, in the form of prognostic and/or predictive biomarkers. This article provides an overview of some of the most recent advances in precision medicine in oncology, with a focus on novel tissue-agnostic anticancer therapies. The definition and implementation of biomarkers and companion diagnostics in clinical trials and clinical practice are also discussed, as well as the changing landscape in clinical trial design.

MRTP: a multiflow real-time transport protocol for ad hoc networks

2006 ◽  
Vol 8 (2) ◽  
pp. 356-369 ◽  
Author(s):  
Shiwen Mao ◽  
D. Bushmitch ◽  
S. Narayanan ◽  
S.S. Panwar
Keyword(s):  
Ad Hoc Networks ◽  
Real Time ◽  
Ad Hoc ◽  
Transport Protocol ◽  
Hoc Networks

Research on the Distributed Characteristics of the Delay for Network Coding Based Real-Time Multicast in MANETs

2014 ◽  
Vol 530-531 ◽  
pp. 768-772
Author(s):  
Guo Ping Tan ◽  
Lin Feng Tan ◽  
Lei Cao ◽  
Mei Yan Ju
Keyword(s):  
Maximum Likelihood ◽  
Probability Distribution ◽  
Network Coding ◽  
Real Time ◽  
Lognormal Distribution ◽  
Ad Hoc ◽  
Estimation Method ◽  
Likelihood Estimation ◽  
Actual Distribution ◽  
Delay Distribution

For the study of the applications of partial network coding based real-time multicast protocol (PNCRM) in Mobile Ad hoc networks, the researches should be developed in the probability distribution of delay. In this paper, NS2 is used to obtain the delay of data packets through simulations. Because the delay does not obey the strict normal distribution, the maximum likelihood estimate method based on the lognormal distribution is used to process the data. Using MATLAB to obtain the actual distribution of the natural logarithm of delay, then drawing the delay distribution with the maximum likelihood estimation method based on the lognormal distribution, the conclusion that the distributions obtained by the above mentioned methods are basically consistent can be obtained. So the delay distribution of PNCRM meets the lognormal distribution and the characteristic of delay probability distribution can be estimated.

Performance Analysis of a Flexible MAC Protocol for Real-Time Services in Vehicular Ad-Hoc Networks

2005 ◽  
Vol 12 (3) ◽  
pp. 147-157
Author(s):  
Giuseppe Caizzone ◽  
Paolo Giacomazzi ◽  
Luigi Musumeci ◽  
Gabriella Saddemi ◽  
Giacomo Verticale
Keyword(s):  
Performance Analysis ◽  
Ad Hoc Networks ◽  
Real Time ◽  
Vehicular Ad Hoc Networks ◽  
Ad Hoc ◽  
Mac Protocol ◽  
Hoc Networks

scholarly journals NtRNF217, Encoding a Putative RBR E3 Ligase Protein of Nicotiana tabacum, Plays an Important Role in the Regulation of Resistance to Ralstonia solanacearum Infection

2021 ◽  
Vol 22 (11) ◽  
pp. 5507
Author(s):  
Ying Liu ◽  
Yuanman Tang ◽  
Xi Tan ◽  
Wei Ding
Keyword(s):  
Nicotiana Tabacum ◽  
Ralstonia Solanacearum ◽  
Plant Immunity ◽  
E3 Ligase ◽  
Marker Genes ◽  
Susceptible Cultivar ◽  
Wild Type ◽  
Disease Symptoms ◽  
Short Time ◽  
Resistance To Ralstonia Solanacearum

E3 ubiquitin ligases, the most important part of the ubiquitination process, participate in various processes of plant immune response. RBR E3 ligase is one of the E3 family members, but its functions in plant immunity are still little known. NtRNF217 is a RBR E3 ligase in tobacco based on the sequence analysis. To assess roles of NtRNF217 in tobacco responding to Ralstonia solanacearum, overexpression experiments in Nicotiana tabacum (Yunyan 87, a susceptible cultivar) were performed. The results illuminated that NtRNF217-overexpressed tobacco significantly reduced multiplication of R. solanacearum and inhibited the development of disease symptoms compared with wild-type plants. The accumulation of H2O2 and O2− in NtRNF217-OE plants was significantly higher than that in WT-Yunyan87 plants after pathogen inoculation. The activities of CAT and SOD also increased rapidly in a short time after R. solanacearum inoculation in NtRNF217-OE plants. What is more, overexpression of NtRNF217 enhanced the transcript levels of defense-related marker genes, such as NtEFE26, NtACC Oxidase, NtHIN1, NtHSR201, and NtSOD1 in NtRNF217-OE plants after R. solanacearum inoculation. The results suggested that NtRNF217 played an important role in regulating the expression of defense-related genes and the antioxidant enzymes, which resulted in resistance to R. solanacearum infection.

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