基因表达差异显示实验

Meta-analysis is an important tool for integrating information from multiple quantitative trait loci (QTLs) studies. Pooling of results from several studies allows greater statistical power for QTL detection and more precise estimation of their genetic effects. Hence, a meta-ana ...

A database can be used to warehouse quantitative trait loci (QTL) data from multiple sources for comparison, genomic data mining, and meta-analysis. A robust database design involves sound data structure logistics, meaningful data transformations, normalization, and proper user i ...

Tremendous progress has been made in recent years on developing statistical methods for mapping quantitative trait loci (QTL) from crosses of inbred lines. In this chapter, we provide an introduction of composite interval mapping and multiple interval mapping methods for mapping QTL f ...

The availability of genetic markers in many species has enabled the analysis of marker-trait associations ranging from small genomic regions to genome-wide scale. An appropriate set of markers must be identified to meet the objectives of any research, using a custom discovery and selecti ...

Genetic mapping methods typically rely upon genotyping many individuals in a mapping population. In contrast, bulk segregant analysis looks for biases in genotype in phenotyped pools of segregants. For relatively strong and genetically simple traits, it can be a fast, inexpensive app ...

Recombinant inbred lines (RILs) are a collection of strains that can be used to map quantitative trait loci. Parent strains are crossed to create recombinants that are then inbred to isogenicity, resulting in a permanent resource for trait mapping and analysis. Here I describe the process of des ...

This chapter covers the procedure of mapping quantitative trait loci (QTLs) in an F2 breeding design. I describe genetic design, general methods and software, and several commonly used approaches. The genetic design section includes F2 population construction. Widely used methods and ...

The development of near isogenic lines (NILs) through repeated backcrossing of genetically distinct parental lines is rather straightforward. Nonetheless, depending on the available resources and the purpose of the lines to be generated, several choices can be made to guide the design ...

The predominate form of DNA diagnostics remains nucleic acid sequencing in the research and clinical setting. While DNA sequencing allows a mutation to be correctly identified, only RNA sequencing can confirm the effect of that mutation on the resulting mRNA transcript. In the absence of RNA ...

Our knowledge about human genes and the consequences of mutations leading to human genetic diseases has drastically improved over the last few years. It has been recognized that many mutations are indeed pathogenic because they impact the mRNA rather than the protein itself. With our better u ...

Yeast is a proven host for the production of recombinant proteins, which may be incorporated in cellular membranes or localized in subcellular compartments. In order to gain access to these proteins, cellular disruption is required to permit extraction, purification, and downstream a ...

Having access to suitably stable, functional recombinant protein samples underpins diverse academic and industrial research efforts to understand the workings of the cell in health and disease. Synthesising a protein in recombinant host cells typically allows the isolation of the ...

The production of recombinant therapeutic proteins is an active area of research in drug development. These bio-therapeutic drugs target nearly 150 disease states and promise to bring better treatments to patients. However, if new bio-therapeutics are to be made more accessible and aff ...

The production of membrane proteins in the large quantities necessary for structural analysis requires many optimization steps. The GFP-fusion-based scheme described in earlier chapters (Chapters 4, 8, and 16) facilitates these steps by allowing the selection of high yielding clon ...

One of the major advantages of using Pichia pastoris is that it is readily adapted to large-scale culture in bioreactors. Bioreactors allow precise regulation of cell growth parameters increasing both yields and reproducibility of the culture. P. pastoris cultures grow to very high cell de ...

Recombinant membrane protein yields can be optimized in Saccharomyces cerevisiae by adjusting the induction time and temperature and/or by the addition of chemical chaperones. Here we describe a protocol for assessing the importance of these parameters.

Pichia pastoris is a widely used host for recombinant protein production. The foaming associated with culturing it on a large scale is commonly prevented by the addition of chemical antifoaming agents or “antifoams.” Unexpectedly, the addition of a range of antifoams to both shake flask and bi ...

The overproduction of eukaryotic membrane proteins in milligram quantities is a major bottleneck for their further biochemical and structural investigation. Production trials exploring a range of input factors can be rationalized to improve the likelihood of success. Here we dis ...

Pichia pastoris is an established host for the production of a wide range of recombinant proteins including membrane proteins. The system has a particularly good track record for the production of G protein-coupled receptors (GPCRs). Generation and screening of expression clones with t ...

There are several ways to introduce non-native DNA into yeast cells, including chemical transformation and electroporation. Methods for both of these procedures are outlined in this chapter. Both methods permit the uptake of DNA from the environment through yeast cell membranes and this ...