生物化学技术

Pseudouridines and 2′-O-methylated nucleotides are ubiquitous constituents of stable cellular RNAs. In eukaryotes, posttranscriptional synthesis of most pseudouridines and 2′-O-methylated nucleotides is directed by sequence-specific guide RNAs (gRNAs). In recent ye ...

Tandem collision-induced dissociation (CID) mass spectrometry (MS) provides a sensitive means of analyzing the amino acid sequence of peptides. Modern MS instrumentation is capable of rapidly generating many thousands of tandem mass spectra, and protein database search engines ...

Proteins are commonly identified through enzymatic digestion and generation of short sequence tags or fingerprints of peptide masses by mass spectrometry. Separation methods, such as liquid chromatography and electrophoresis, are often used to fractionate complex protein or ...

The goal of quantitative proteomics is to systematically study static state or perturbation-induced changes in protein profile. Most of the recently developed mass spectrometry (MS)-based quantitative proteomic methods employ stable isotope labeling to introduce signatu ...

Difference gel electrophoresis (DIGE) technology provides a powerful quantitative component to proteomics experiments involving two-dimensional (2D) gel electrophoresis. DIGE allows for the detection of subtle changes in protein abundance with statistical confiden ...

Proteios (http://www.proteios.org) is an initiative for the development of a comprehensive open source system for storage, organization, analysis, and annotation of proteomics experiments. The Proteios platform is based on existing principles for proteomics data publishing a ...

The ever increasing volumes of proteomic data now being produced by laboratories across the world have resulted in major issues in data storage and accessibility. The further demands of multilaboratory initiatives has highlighted issues when collaborators cannot import data gen ...

In recent years the organization of mass spectrometry (MS) data obtained in large-scale proteomics projects became an important issue. This has catalyzed the development of a few different database schemes for storing MS data, as well as some dedicated user interfaces. However, many of these ...

The analysis method described in this chapter demonstrates the structural characterization of carbohydrates based on their molecular mass, as well as the mass of their respective fragment ions using mass spectrometry (MS). The carbohydrate molecules are first converted into gase ...

The intention with this chapter is to give an overview of a broad range of freely available programs on the internet which are useful for analyses of mass spectrometry data. The list is by no means a full list of free proteomics tools on the net and I apologize if there are other good tools on the internet that have be ...

Mass spectrometry (MS) has recently become one of the most informative methods for studying proteins. Albiet, MS cannot compete with the detailed structural information obtained by methods such as nuclear magnetic resonance and X-ray crystallography. However, MS is much easier to auto ...

Peak extraction from raw data is the first step in analysis of mass spectrometry (MS) data. The quality of this procedure is very important because it affects the quality of all subsequent analysis, such as database searches and peak quantitation. Many methods have been proposed in the literatur ...

With the introduction of simple and relatively inexpensive methods for labeling nucleic acids with nonradioactive labels, doors have been opened that enable nucleic acid hybridization probes to be used for research and development, as well as for clinical diagnostic applications. ...

Fluorescent energy transfer within dual-labeled oligonucleotide probes is widely used in assays for genetic analysis. Nucleic acid detection/amplification methods, such as real-time polymerase chain reaction, use dual-labeled probes to measure the presence and copy number ...

Induced fluorescence resonance energy transfer (iFRET) is a variation of resonance energy transfer that is particularly well-suited for the detection of DNA hybridization. The underlying mechanism involves monitoring changes in fluorescence that are the result of an energy tra ...

Fluorescence resonance energy transfer (FRET) occurs when two fluorophores are in close proximity, and the emission energy of a donor fluorophore is transferred to excite an acceptor fluorophore. Using such fluorescently labeled oligonucleotides as FRET probes, makes possible s ...

Several types of DNA cuts are used as markers of apoptosis for detection of apoptotic cells in situ. We recently introduced a ligase-based in situ assay that is specific for a single type of DNA damage—a double-strand break of DNase I-type, bearing 5′PO4. Here we describe a vaccinia topoisomerase I-ba ...

A novel method for DNA enzymatic cleavage assays using molecular beacons (MBs) as the substrate for nuclease is described. An MB is a hairpin-shaped DNA probe that is labeled with a fluorescent dye at one end and a quencher at the other end. The loop sequence of the MB can be used as the substrate for single-stra ...

Exploring the properties of molecules that cleave DNA (i.e., enzymatic nucleases, chemical footprinting agents, and naturally occurring DNA cleaving antibiotics) has been an ongoing process with benefits extending toward both laboratory and clinical applications. Despite t ...

Quantitative polymerase chain reaction assays have become the most common methods in the determination of viral load during cytomegalovirus (CMV) infection of transplant patients. In recent years, the development of automated nucleic acid extraction devices together with the i ...