遗传学实验技术

Small-molecule drugs are a cost-effective way to treat and prevent disease. A study by the Slone Institute published in 2002 estimated that over 50% of the adult population in the United States used at least one pharmaceutical drug during the preceding week. The positive impact of small-molecule ...

New methods and strategies have been developed to design and use small molecules that allow the functional dissection of molecular pathways, cells, and organisms by selective small-molecule ligands or modulators. In this overview, we are focusing on diversity aspects, design methods, ...

In the current post-genomic era, cell-free translation platforms are gaining importance in structural as well as functional genomics. They are based on extracts prepared from Escherichia coli cells, wheat germ, or rabbit reticulocytes, and when programmed with any mRNA in the presence of ...

This chapter contains protocols for high-throughput protein production based on the cell-free system prepared from eukaryote wheat embryos.

De novo design provides an in silico toolkit for the design of novel molecular structures to a set of specified structural constraints, and is thus ideally suited for creating molecules for chemical genomics. The design process involves manipulation of the input, modification of structu ...

Recombinant adenoviruses are versatile tools for gene delivery into mammalian tissue-culture cells. Adenoviruses can infect both dividing and nondividing cells, with efficiencies approaching 100%. Here we describe the use of adenoviruses to express reporter proteins in high- ...

This chapter describes protocols for preparing 15N-labeled proteins (ubiquitin is used as an example) using Escherichia coli cells (with purification) and the wheat germ cell-free system (without purification). A comparison of 1H-15N heteronuclear single-quantum coherence ( ...

Protein microarrays have many potential applications in high-throughput analysis of protein function. However, simple, reproducible, and robust methods for array fabrication are required. Here we discuss the background to different routes to array fabrication and describe in d ...

Matrix-assisted laser desorption/ionization (MALDI)-time-of-flight (TOF)-mass spectrometry (MS) is now routinely used in many laboratories for the rapid and sensitive identification of proteins by peptide mass fingerprinting (PMF). We describe a simple protocol that can be p ...

We have established a high-throughput biochip platform for constructing carbohydrate microarrays. Using this technology, carbohydrate-containing macromolecules of diverse structures, including polysaccharides, natural glycoconjugates, and mono- and oligosa ...

A yeast two-hybrid screening system was developed to screen for small molecules that inhibit the interaction of the Ras and the Raf proteins. Hyperpermeable yeast strains useful for high-throughput screening (HTS) for the two-hybrid system were created. Differential inhibition of the ...

Affinity chromatography is ideally suited to the purification of pharmaceutical proteins due to its unique bio-specificity characteristics. Tailor-made affinity ligands that represent a promising class of synthetic affinity ligands have been developed to target specific ...

In silico docking techniques are being used to investigate the complementarity at the molecular level of a ligand and a protein target. As such, docking studies can be used to identify the structural features that are important for binding and for in silico screening efforts in which suitable bin ...

The lipopolysaccharide of capsule-deficient Haemophilus infuenzae strain Rd contains an N-acetylgalactosamine residue attached to the terminal globotriose moiety in the Hex5 glycoform. Genome analysis identified an open reading frame, HI1578, referred to as LgtD, whose ami ...

Protein expression and purification have traditionally been time-consuming, case-specific endeavors, and are considered to be the greatest bottlenecks in most proteomics pipelines. Escherichia coli (E. coli) is the most convenient and cost-effective host, although optimal c ...

Ligation-independent cloning (LIC) is a simple, rapid, and efficient method for high-throughput cloning. In this system, linear plasmid vector and insert DNA are treated to generate complementary single-stranded overhangs that anneal during a short incubation. The LIC system is ada ...

This chapter describes a method for efficient high-throughput purification of hexahistidine-tagged proteins that are expressed in Escherichia coli (E. coli) using immobilized metal affinity chromatography (IMAC) (2) in a 96-well format. This approach is particularly suitable ...

The common cardiovascular, respiratory, gastrointestinal tract, renal, and immune diseases share common etiologies, because they are multifactorial in origin, with both genetic and environmental components contributing to their clinical phenotype. Recently, the necess ...

Analysis of differential antigen expression in tumor and metastases vs normal tissue is frequently performed in oncological research. If suitable antibodies are available, its specificity is tested in Western blot, and the distribution of the antigen in the tissue is followed by immun ...

The human genome contains perhaps more than 50,000 genes. A typical cell in the human body is believed to express from 15,000 to 25,000 proteins. The majority of these proteins are expressed in all the cells, and are known as housekeeping genes. These are proteins involved in vital cell functions, inclu ...