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Rescooped by Rebecca McDougal from The science toolbox
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PhySortR: a fast, flexible tool for sorting phylogenetic trees in R

PhySortR: a fast, flexible tool for sorting phylogenetic trees in R | molecular tools | Scoop.it
A frequent bottleneck in interpreting phylogenomic output is the need to screen often thousands of trees for features of interest, particularly robust clades of specific taxa, as evidence of monophyletic relationship and/or reticulated evolution. Here we present PhySortR, a fast, flexible R package for classifying phylogenetic trees. Unlike existing utilities, PhySortR allows for identification of both exclusive and non-exclusive clades uniting the target taxa based on tip labels (i.e., leaves) on a tree, with customisable options to assess clades within the context of the whole tree. Using simulated and empirical datasets, we demonstrate the potential and scalability of PhySortR in analysis of thousands of phylogenetic trees without a priori assumption of tree-rooting, and in yielding readily interpretable trees that unambiguously satisfy the query. PhySortR is a command-line tool that is freely available and easily automatable.

Via Niklaus Grunwald
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Rescooped by Rebecca McDougal from Plants and Microbes
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bioRxiv: Efficient Disruption and Replacement of an Effector Gene in the Oomycete Phytophthora sojae using CRISPR/Cas9 (2015)

bioRxiv: Efficient Disruption and Replacement of an Effector Gene in the Oomycete Phytophthora sojae using CRISPR/Cas9 (2015) | molecular tools | Scoop.it

Phytophthora sojae is a pathogenic oomycete that infects soybean seedlings as well as stems and roots of established plants, costing growers $1–2 billion per year. Due to its economic importance, P. sojae has become a model for the study of oomycete genetics, physiology and pathology. Despite the availability of several genome sequences, the lack of efficient techniques for targeted mutagenesis and gene replacement have long hampered genetic studies of pathogenicity in Phytophthora species. Here, we describe a CRISPR/Cas9 system enabling rapid and efficient genome editing in P. sojae. Using the RXLR effector gene Avr4/6 as target, we observed that in the absence of a homologous template, the repair of Cas9-induced double-strand breaks (DSBs) in P. sojae was mediated by non-homologous end joining (NHEJ), primarily resulting in short indels. Most mutants were homozygous, presumably due to gene conversion triggered by Cas9-mediated cleavage of non-mutant alleles. When donor DNA was present, homology directed repair (HDR) was observed, which resulted in the replacement of the target gene with the donor DNA. By testing the specific virulence of several NHEJ mutants and HDR -mediated gene replacements on soybeans, we have validated the contribution of Avr4/6 to recognition by soybean R gene loci, Rps4 and Rps6, but also uncovered additional contributions to resistance by these two loci. Our results establish a powerful tool for studying functional genomics in Phytophthora, which provides new avenues for better control of this pathogen.


Via Kamoun Lab @ TSL
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Rescooped by Rebecca McDougal from The science toolbox
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Speeding Up Ecological and Evolutionary Computations in R; Essentials of High Performance Computing for Biologists

Speeding Up Ecological and Evolutionary Computations in R; Essentials of High Performance Computing for Biologists | molecular tools | Scoop.it
Computation has become a critical component of research in biology. A risk has emerged that computational and programming challenges may limit research scope, depth, and quality. We review various solutions to common computational efficiency problems in ecological and evolutionary research. Our review pulls together material that is currently scattered across many sources and emphasizes those techniques that are especially effective for typical ecological and environmental problems. We demonstr

Via Niklaus Grunwald
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Rescooped by Rebecca McDougal from Diagnostic activities for plant pests
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Affordable and reliable plant sap-mediated template preparation for the detection of various phytopathogens by PCR assay - Sahana et al. 2014

Affordable and reliable plant sap-mediated template preparation for the detection of various phytopathogens by PCR assay - Sahana et al. 2014 | molecular tools | Scoop.it

Plant pathogens like ‘Ca. Liberibacter’, phytoplasma, viruses and viroids cause diseases to almost all economically important plants. A simple and fast sap-mediated polymerase chain reaction (PCR) method for the detection of these pathogens infecting various plant hosts is described in the present study. Sap from selected plants was drawn aseptically on parafilm, from the mid-rib of young leaves. Depending on the type of host plant, sap was diluted to optimal concentration before PCR analysis. ‘Ca. Liberibacter’, Citrus tristeza virus (CTV) and Citrus exocortis viroid (CEVd) infecting citrus, and ‘Ca. Phytoplasma’ infecting pepper and sandal trees were tested by sap-mediated PCR. The reliability of this procedure was evaluated by comparing the findings with previously described protocols. The sap-mediated nucleic acid template preparation for PCR assay is devoid of laborious nucleic acid extraction and expensive chemicals. Hence the present method is rapid, economical and so can be employed for diagnosis of large number of plant samples


Via Petter Françoise
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Rescooped by Rebecca McDougal from Plant Pathogenomics
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PLOS Pathogens: Mining Herbaria for Plant Pathogen Genomes: Back to the Future (2014)

PLOS Pathogens: Mining Herbaria for Plant Pathogen Genomes: Back to the Future (2014) | molecular tools | Scoop.it

Since the dawn of agriculture, plant pathogens and pests have been a scourge of humanity. Yet we have come a long way since the Romans attempted to mitigate the effects of plant disease by worshipping and honoring the god Robigus. Books in the Middle Ages by Islamic and European scholars described various plant diseases and even proposed particular disease management strategies. Surprisingly, the causes of plant diseases remained a matter of debate over a long period. It took Henri-Louis Duhamel du Monceau's elegant demonstration in his 1728 “Explication Physique” paper that a “contagious” fungus was responsible for a saffron crocus disease to usher in an era of documented scientific inquiry. Confusion and debate about the exact nature of the causal agents of plant diseases continued until the 19th century, which not only saw the first detailed analyses of plant pathogens but also provided much-needed insight into the mechanisms of plant disease. An example of this is Anton de Bary's demonstration that a “fungus” is a cause, not a consequence, of plant disease. This coming of age of plant pathology was timely. In the 19th century, severe plant disease epidemics hit Europe and caused economic and social upheaval. These epidemics were not only widely covered in the press but also recognized as serious political issues by governments. Many of the diseases, including late blight of potato, powdery and downy mildew of grapevine, as well as phylloxera, were due to exotic introductions from the Americas and elsewhere. These and subsequent epidemics motivated scientific investigations into crop breeding and plant disease management that developed into modern plant pathology science over the 20th century.

 

Nowadays, our understanding of plant pathogens and the diseases they cause greatly benefits from molecular genetics and genomics. All aspects of plant pathology, from population biology and epidemiology to mechanistic research, are impacted. The polymerase chain reaction (PCR) first enabled access to plant pathogen DNA sequences from historical specimens deposited in herbaria. Historical records in combination with herbarium specimens have turned out to provide powerful tools for understanding the course of past plant epidemics. Recently, thanks to new developments in DNA sequencing technology, it has become possible to reconstruct the genomes of plant pathogens in herbaria. In this article, we first summarize how whole genome analysis of ancient DNA has been recently used to reconstruct the 19th-century potato-blight epidemic that rapidly spread throughout Europe and triggered the Irish potato famine. We then discuss the exciting prospects offered by the emergence of the discipline of ancient plant pathogen genomics.


Via Kamoun Lab @ TSL
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Mary Williams's curator insight, April 25, 2014 3:02 AM

Good overview for students - very accessible and interesting!

Freddy Monteiro's comment, April 25, 2014 4:21 AM
This is a great source of teaching materials for potato late blight. Congrats on the work behind it.
Rescooped by Rebecca McDougal from Pharma Biotech Industry Review (Krishan Maggon)
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Choosing the Right Tool for the Job: RNAi, TALEN, or CRISPR: Molecular Cell

Choosing the Right Tool for the Job: RNAi, TALEN, or CRISPR: Molecular Cell | molecular tools | Scoop.it

Molecular Cell Abstract

The most widely used approach for defining gene function is to reduce or completely disrupt its normal expression. For over a decade, RNAi has ruled the lab, offering a magic bullet to disrupt gene expression in many organisms. However, new biotechnological tools—specifically CRISPR-based technologies—have become available and are squeezing out RNAi dominance in mammalian cell studies. These seemingly competing technologies leave research investigators with the question: “Which technology should I use in my experiment?” This review offers a practical resource to compare and contrast these technologies, guiding the investigator when and where to use this fantastic array of powerful tools.


Via Krishan Maggon
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Krishan Maggon 's curator insight, May 26, 2015 7:28 AM

Molecular Cell

 

Volume 58, Issue 4, p575–585, 21 May 2015Review Choosing the Right Tool for the Job: RNAi, TALEN, or CRISPRMichael Boettcher, Michael T. McManus DOI: http://dx.doi.org/10.1016/j.molcel.2015.04.028 |  ;
Rescooped by Rebecca McDougal from The science toolbox
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The Quantitative Methods Boot Camp: Teaching Quantitative Thinking and Computing Skills to Graduate Students in the Life Sciences

The Quantitative Methods Boot Camp: Teaching Quantitative Thinking and Computing Skills to Graduate Students in the Life Sciences | molecular tools | Scoop.it
The past decade has seen a rapid increase in the ability of biologists to collect large amounts of data. It is therefore vital that research biologists acquire the necessary skills during their training to visualize, analyze, and interpret such data. To begin to meet this need, we have developed a “boot camp” in quantitative methods for biology graduate students at Harvard Medical School. The goal of this short, intensive course is to enable students to use computational tools to visualize and

Via Niklaus Grunwald
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Rescooped by Rebecca McDougal from Diagnostic activities for plant pests
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EPPO Workshop on setting Ct cut-off values for real-time PCR Paris, 2013-11-11/12

EPPO Workshop on setting Ct cut-off values for real-time PCR Paris, 2013-11-11/12 | molecular tools | Scoop.it

Read the conclusions of the Workshop


Via Petter Françoise
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Petter Françoise's curator insight, November 26, 2013 11:29 AM

The Workshop allowed the improvement of the intructions to authors of EPPO Diagnostic Protocols. 

Rescooped by Rebecca McDougal from Diagnostic activities for plant pests
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Optimising droplet digital PCR analysis approaches for detection and quantification of bacteria: a case study of fire blight and potato brown rot -Dreo et al 2014

Optimising droplet digital PCR analysis approaches for detection and quantification of bacteria: a case study of fire blight and potato brown rot -Dreo et al 2014 | molecular tools | Scoop.it

Here we report on the first assessment of droplet digital PCR (ddPCR) for detection and absolute quantification of two quarantine plant pathogenic bacteria that infect many species of the Rosaceae and Solanaceae families: Erwinia amylovora and Ralstonia solanacearum. An open-source R script was written for the ddPCR data analysis. Analysis of a set of samples with known health status aided the assessment and selection of different threshold settings (QuantaSoft analysis, definetherain pipeline and manual threshold), which led to optimal diagnostic specificity. The interpretation of theE. amylovora ddPCR was straightforward, and the analysis approach had little influence on the final results and the concentrations determined. The sensitivity and linear range were similar to those for real-time PCR (qPCR), for the analysis of both bacterial suspensions and plant material, making ddPCR a viable choice when both detection and quantification are desired. With the R. solanacearum ddPCR, the use of a high global threshold was necessary to exclude false-positive reactions that are sometimes observed in healthy plant material. ddPCR significantly improved the analytical sensitivity over that of qPCR, and improved the detection of low concentrations of R. solanacearum in potato tuber samples. Accurate and rapid absolute quantification of both of these bacteria in pure culture was achieved by direct ddPCR. Our data confirm the suitability of these ddPCR assays for routine detection and quantification of plant pathogens and for preparation of defined in-house reference materials with known target concentrations.


Via Petter Françoise
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BioTechniques - MIQE qPCR Guidelines, Three Years Later

BioTechniques - MIQE qPCR Guidelines, Three Years Later | molecular tools | Scoop.it
Three years ago, a group of researchers began campaigning for the adoption of guidelines that promised to improve the reproducibility of RT-qPCR data. Lauren Arcuri Ware reports on the adoption and evolution of these guidelines....
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