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Fluorescence in Situ Hybridization Techniques for Cytogenetic and Genomic Analyses
Time of Update: 2021-01-17
Yet despite the value of FISH analysis for cytogenetic studies, there are surprisingly few labs that are able to adapt the technique for their experiments in chromosomal and genome biology.
Our description uses rice as a model for performing a complete FISH procedure, but the protocol can be readily adapted for other plant species.
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Activation Tagging and Insertional Mutagenesis in Barley
Time of Update: 2021-01-17
This has been achieved primarily by high-throughput plant transformation using T- DNA sequences that encode regulatory elements.
Insertion of this transposable element into genes can also generate insertional inactivation mutants enabling both gene overexpression and gene knockout mutants to be identified in the same population.
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Citrus
Time of Update: 2021-01-17
Abundant availability of starting material and relative simplicity make this procedure an attractive choice for many researchers despite transformation efficiency that is in the low range of about 1%.
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The Quantitative Real-Time Polymerase Chain Reaction for the Analysis of Plant Gene Expression
Time of Update: 2021-01-17
The quantitative real-time polymerase chain reaction is used to simultaneously amplify and quantify a targetedDNA molecule.
When combined with reverse transcription, it is a powerful tool for the analysis of gene expression, and it is widely used for this purpose in plant species.
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Begomoviruses: Molecular Cloning and Identification of Replication Origin
Time of Update: 2021-01-17
TheBegomovirus genus is the largest genus of theGeminiviridae family and comprises the whitefly transmitted geminiviruses that infect dicotyledonous plants.
In this chapter, we describe the cloning of begomovirus replication modules and the subsequent functional characterization of geminivirus replication origins.
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Systems Approaches to Unraveling Plant Metabolism: Identifying Biosynthetic Genes of Secondary Metabolic Pathways
Time of Update: 2021-01-17
Cimicifuga racemosa , Nutt., black coshosh), a non-model medicinal plant with no genome sequence and little horticultural information available, that have led to the development of initial gene–metabolite relationships for the production of several bioactive metabolites in this multicomponent botanical therapeutic, and that can be readily applied to a wide variety of under-characterized medicinal plants.
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Screening Chemical Libraries for Compounds That Affect Protein Sorting to the Yeast Vacuole
Time of Update: 2021-01-17
In these genetic screens, mis-targeted vacuolar cargo proteins were used as phenotype markers.
Here we describe a similar approach employing pharmacological effects of diverse chemical compounds to mimic molecular phenotypes caused by conventional genetic mutations in protein trafficking genes.
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A Floral Induction System for the Study of Early Arabidopsis Flower Development
Time of Update: 2021-01-17
This is especially problematic when investigating molecular events at very early stages ofArabidopsis flower development, as the floral buds are minute and are initiated sequentially such that a single flower on an inflorescence is at a given developmental stage.
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Analysis of Population Structures of Viral Isolates using Single StrandConformation Polymorphism (SSCP) Method
Time of Update: 2021-01-17
The single-strand conformation polymorphism (SSCP) makes possible the identification of genetic differences between viral sequences and constitutes an alternative to the expensive and time-consuming cloning and sequencing strategies.
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Seed Proteomics
Time of Update: 2021-01-17
Rather than providing a single specific protocol, the inclusive area of seed proteomics is reviewed; methods are described and compared and primary literature citations are provided.
The proteomic analysis of seeds encounters some specific problems that do not impinge on analyses of other plant cells, tissues, or organs.
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Trichome Development in Arabidopsis
Time of Update: 2021-01-17
Arabidopsis trichomes are giant single epidermal cells that are easily accessible for genetic, genomic and cell-biological analysis.
Trichome studies are greatly facilitated by methods specifically applicable for this particular cell type.
This chapter will highlight these two aspects of trichome analysis.
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Orchids (Cymbidium spp., Oncidium, and Phalaenopsis)
Time of Update: 2021-01-17
This chapter presents a simple and reproducibleAgrobacterium tumefaciens -mediated transformation protocol and molecular screening technique of transgenics for two orchid species,Oncidium andPhalaenopsis .
Different molecular methods and GUS-staining used to screen putative transgenic plants to confirm the integration of foreignDNA into the orchid genome were also described in detail.
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Pull-Down Assays for Plant Hormone Research
Time of Update: 2021-01-17
Hormonal signals are transduced (and sometimes perceived) by protein–protein interactions.
Understanding these interactions is therefore crucial to understanding the network of signalling components as a whole.
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Manipulating Large-Scale Arabidopsis Microarray Expression Data: Identifying Dominant Expression Patterns and Biological Process Enrichment
Time of Update: 2021-01-17
Therefore, finding these co-expressed groups can reduce the dimensionality of complex expression data into a set of dominant transcriptional regulatory modules.
Using these new methods we can begin to understand the biological information contained within large-scale expression data sets.
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Visualizing Auxin Transport Routes in Arabidopsis Leaf Primordia
Time of Update: 2021-01-17
Visualization of apparent auxin transport patterns has largely been facilitated by the recent creation of translational fusions of GFP to members of theArabidopsis (At)PIN family of auxin efflux associated proteins.
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Detection of mRNA Expression Patterns by Nonradioactive In Situ Hybridization on Histological Sections of Floral Tissue
Time of Update: 2021-01-17
Here, we present a robust protocol for sensitive analysis of expression patterns in inflorescence tissue ofArabidopsis thaliana .
While the described protocol is optimized for inflorescence meristems, it can possibly be used for other tissues as well.
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Array Platforms and Bioinformatics Tools for the Analysis of Plant Transcriptome in Response to Abiotic Stress
Time of Update: 2021-01-17
Current microarray technologies allow high-density in situ synthesis of oligonucleotides or ex situ spotting of target molecules (c DNA ) for conducting genome-wide comparative gene expression profiling studies.
In this chapter we summarize the technical issues of different microarray technologies, discuss the availability of bioinformatics tools, and present approaches to extract biologically meaningful knowledge.
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Phosphorylation Analysis of Plant Viral Proteins
Time of Update: 2021-01-17
This chapter outlines the concepts and methods used to investigate protein phosphorylation and its physiological relevance during plant virus infection.
Rather than providing an exhaustive review of the experimental protocols for protein phosphorylation analysis, we focus on methods that can be used to study phosphorylation of viral proteins.
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Japonica Rice Varieties (Oryza sativa, Nipponbare, and Others)
Time of Update: 2021-01-17
Several protocols have been developed and fine-tuned for particular genotypes, including commercial genotypes, making use of either mature seeds or immature embryos as target tissue forAgrobacterium infection.
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Isolation of Proteins Binding to Promoter Elements of Alkaloid Metabolism-Related Genes Using Yeast One-Hybrid
Time of Update: 2021-01-16
Transcription factors involved in biosynthetic regulation can be isolated based on their ability to bind these specific promoter elements using yeast one-hybrid screening.
The aim of this chapter is to describe the yeast one-hybrid system for screening DNA-binding proteins potentially involved in transcriptional regulation.
