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Identification of Plant microRNAs Using Expressed Sequence Tag Analysis
Time of Update: 2020-11-18
EST analysis is based on the conservation of miRNA sequences and the stem-loop hairpin secondary structures of miRNAs. In this method, potential miRNAs will first be identified by EST analysis followed by confirmation using TaqMan � MicroRNA qRT- PCR .
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In Vitro Culture of Arabidopsis Embryos
Time of Update: 2020-11-18
However, experimental manipulation with embryogenesis is difficult, as the embryo develops deeply inside maternal tissues.
It allows pharmacological manipulation of embryo development and does not interfere with standard techniques for localizing gene expression and protein localization in the cultivated embryos.
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Agrobacterium-Mediated Transformation of Potato Genotypes
Time of Update: 2020-11-18
The potato is ideally suited for improvement by modern genetic manipulation methods, as it is highly amenable to a wide range of various tissue-culture techniques ( 1 ).
Genetic transformation, however, provides a more direct and controlled method for manipulating the genome of potato for both basic and applied research purposes ( 3 ).
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Appendix A: The Components of the Culture Media
Time of Update: 2020-11-18
The success in the technology and application of plant tissue culture is greatly influenced by the nature of the culture medium used.
A better understanding of the nutritional requirements of cultured cells and tissues can help to choose the most appropriate culture medium for the explant used.
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DNA-Based Diagnostics for Genetically Modified Cotton: Decaplex PCR Assay to Differentiate MON531 and MON15985 Bt Cotton Events
Time of Update: 2020-11-18
With the increase in development and commercialization of GM crops, it is necessary to develop appropriate qualitative and quantitative methods for detection of different GM events.
This chapter summarizes the methods based on polymerase chain reaction ( PCR ) being employed for detection of different GM events of cotton.
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Lily.
Time of Update: 2020-11-18
lily is available for medicinal use in the Chinese Han Dynasty's Book of Shen Nong Herbs.
From the beginning of the 17th century to the 18th century, lily of American and Chinese origin was introduced to Europe.
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Confocal Microscopy in PlantPathogen Interactions
Time of Update: 2020-11-18
The development of confocal microscopy and its application to studies of plant–pathogen interactions have revolutionised research into the role of selected molecules and cell components in pathogen infection strategies and plant defence responses.
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Isolation and Biochemical Analysis of Plant Small RNAs
Time of Update: 2020-11-18
Further, protocols are given for the analysis and quantification of small RNAs using polyacrylamide gel-based approaches.
A native streptavidin gel-shift assay, useful for measuring the relative amounts of multiple small RNAs simultaneously, is presented.
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Investigating the Phytohormone Ethylene Response Pathway by Chemical Genetics
Time of Update: 2020-11-18
As an alternative approach, chemical screening with natural products or synthetic small molecules that act as conditional mutagens allows for identifying bioactive compounds as bioprobes to overcome the above-mentioned issues.
Here, we describe our methodology for using chemical genetics to identify small molecules that interfere with the ethylene response.
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Hormone-Binding Assay Using Living Bacteria Expressing Eukaryotic Receptors
Time of Update: 2020-11-18
The method includes procedures of bacteria growing, incubation with labeled hormone, separation of bound from unbound ligand, determination of radioactivity in bacterial precipitates, and mathematical analysis of primary data.
The established simple protocol for specific labeling hormone-binding sites in intact bacteria allows determination of the main parameters of the ligand–receptor interaction.
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Computational Tools for Quantitative Analysis of Cell Growth Patterns and Morphogenesis in Actively Developing Plant Stem Cell Niches
Time of Update: 2020-11-18
Therefore, a quantitative and dynamic understanding of the spatial and temporal organization of gene expression and cell behavioral patterns within multilayered and actively growing developmental fields is crucial to model the process of development.
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In Vitro Transcription and Translation
Time of Update: 2020-11-18
These contain one or more promoter sequences, recognized by the T3, T7, or SP6 RNA polymerase enzymes, flanking the multiple-cloning site.
Once cloned, an insert, e.g., a DNA sequence coding for a viral coat protein (CP), may be transcribed into mRNA in a simple reaction exploiting these enzymes.
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The Determination and Quantification of Photosynthetic Pigments by Reverse Phase High-Performance Liquid Chromatography, Thin-Layer Chromatography, and Spectrophotometry
Time of Update: 2020-11-18
Methods for the determination of chlorophyllsa andb , β-carotene, neoxanthin, and the pigments that are involved in photoprotective cycles such as the xanthophylls are discussed.
In addition, several spectrophotometric methods for the quantification of chlorophyllsa andb are described.
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Cowpea Mosaic Virus-Based Systems for the Expression of Antigens and Antibodies in Plants
Time of Update: 2020-11-18
The systems include vectors based on both full-length and deleted versions of RNA-2.
Both types of vector have proved effective for the production in plants of a diverse range of proteins including antigens and antibodies.
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Visualization of Phytoplasmas Using Electron Microscopy
Time of Update: 2020-11-18
The use of electron microscopy, both transmission and scanning, provides reliable and accurate methods for detecting phytoplasmas in plants.
Our understanding of these pathogens, their morphology, development, and intracellular location in plants and insect vectors has been greatly increased through the use of these instruments.
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Affinity Purification and Determination of Enzymatic Activity of Recombinantly Expressed Aldehyde Dehydrogenases
Time of Update: 2020-11-18
Here we describe a straightforward protocol for purification of soluble recombinantly expressed ALDH enzyme based on metal affinity chromatography and the subsequent determination of enzymatic activity using aldehydic substrates, which is assayed spectrophotometrically at 340 nm by conversion of NAD(P)+ to NAD(P)H.
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Phenotypic and Physiological Evaluation for Drought and Salinity Stress Responses in Rice
Time of Update: 2020-11-18
The growing need to improve rice cultivars for drought and salt tolerance requires the development of reproducible screening methods that simulate field conditions, and which provide quantitative data for statistical testing and selection of genotypes with differential responses.
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Knocking Out the Wall: Protocols for Gene Targeting in Physcomitrella patens
Time of Update: 2020-11-18
The mossPhyscomitrella patens has become established as a model for investigating plant gene function due to the feasibility of gene targeting.
patens genome have identified genes that putatively encode cell wall biosynthetic enzymes, providing a basis for investigating the evolution of cell wall polysaccharides and the enzymes that synthesize them.
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Profiling of Hormones and Related Metabolites in Seed Dormancy and Germination Studies
Time of Update: 2020-11-18
Seed dormancy and germination are regulated by several plant hormones, such as abscisic acid, gibberellin, auxin (indole-3-acetic acid), ethylene, and brassinosteroid.
Endogenous concentrations of a hormone are determined by the balance between biosynthesis and deactivation, and contribute to the regulation of physiological responses.
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Thylakoid Phosphoproteins: Identification of Phosphorylation Sites
Time of Update: 2020-11-18
Various methods for the detection of thylakoid phosphoproteins, including the preparation of phosphopeptides for mass spectrometric analyses and techniques for phosphopeptide identification by electrospray ionization mass spectrometry (ESI-MS) are described.
