遗传学实验技术

Small interfering RNAs (siRNAs) have been widely used for knocking down gene expression in a variety of organisms. Although experiments in cancer cell lines indicate that siRNAs are usually not detected by innate immunity, lipid-mediated delivery of siRNAs into blood cells is often accom ...

RNA interference (RNAi) is a natural mechanism for gene silencing that can be harnessed for the development of RNA-based drugs. Although synthetic small interfering RNA (siRNAs) can be delivered in vitro to virtually all cell types using lipid-based transfection agents or electropora ...

RNA interference (RNAi) is a relatively new found phenomenon of posttranscriptional gene silencing to regulate the expression of multiple genes involved in a wide range of biological processes. The gene-silencing technology via RNAi has also been developed into a commonly anti-gene m ...

The fast developing field of RNA interference (RNAi) requires monitoring of small interfering RNA (siRNA) delivery to targeted organs and evaluating the efficiency of target gene silencing. The molecular imaging approach fits perfectly to fulfill these needs and provides informa ...

This chapter describes how to design and conduct experiments to deliver siRNA to adherent mammalian cells in vitro by magnetic force–assisted transfection using self-assembled complexes of small interfering RNA (siRNA) and cationic lipids or polymers that are associated with mag ...

RNA interference (RNAi) is a potent and specific mechanism for eliminating the mRNA of specific genes. This gene silencing mechanism occurs naturally and is highly conserved from plants to human cells, holding promise for functional genomics and for revolutionizing medicine due to its u ...

RNAi is a powerful research tool for specific gene silencing and may also lead to promising novel therapeutic strategies. However, the development of RNAi-based therapies has been slow due to the lack of targeted delivery methods. The biggest challenge in the use of siRNA-based therapies is de ...

Post-transcriptional gene silencing mediated by double-stranded RNA represents an evolutionarily conserved cellular mechanism. Small dsRNAs, such as microRNAs (miRNAs), are part of the main regulatory mechanisms of gene expression in cells. The possibilities of harnessing ...

Bacterial transformation is an essential component of many molecular biological techniques, but bacterial restriction-modification (R-M) systems can preclude the efficient introduction of shuttle vector plasmids into target bacterial cells. Whole-genome DNA sequenc ...

A genetic tool for introducing marker-free deletions is essential for multiple manipulations of genomes. We have developed a simple and efficient method for creating marker-free deletion mutants of Bacillus subtilis through transformation with recombinant PCR products, using ...

This chapter provides methods and insights into the use of broad-host-range plasmid vectors useful for expression of genes in a variety of bacteria. The main focus is on IncQ, IncW, IncP, and pBBR1-based plasmids which have all been used for such applications. The specific design of each vector is ada ...

Reverse genetic approaches have become invaluable tools to tap into the wealth of information provided by sequenced genomes. In 2007, sequencing of the Chlamydomonas reinhardtii genome was completed, and with this an increased demand for the development of reverse genetic strategi ...

Filamentous fungi have received attentions as hosts for heterologous protein production because of their high secretion capability and eukaryotic post-translational modifications. One of the safest hosts for heterologous protein production is Koji mold Aspergillus oryz ...

Thermoacidophilic archaea comprise one of the major classes of extremophiles. Most belong to the family Sulfolobales within the phylum Crenarchaeota. They are of applied interest as sources of hyperstable enzymes, for biomining of base and precious metals, and for evolutionary stud ...

Zymomonas mobilis is a facultatively anaerobic α-proteobacterium with a considerable potential for industrial ethanol production. An important tool in the generation of stable mutants for this organism is described in this chapter; it entails insertional mutagenesis with the h ...

Construction of gene disruption mutants and analysis of the resultant phenotypes are an important strategy to study gene function. A simple and high-throughput method developed for microorganisms combines two different types of transposons, direct genomic DNA amplification a ...

The genus Clostridium is a diverse assemblage of Gram positive, anaerobic, endospore-forming bacteria. Whilst certain species have achieved notoriety as important animal and human pathogens (e.g. Clostridium difficile, Clostridium botulinum, Clostridium tetani, and Clos ...

Lactobacillus acidophilus NCFM is a probiotic microbe with the ability to survive passage to the �gastrointestinal tract, interact intimately with the host and induce immune responses. The genome of NCFM has been determined and the bacterium is genetically accessible. Therefore, L. a ...

The depth of knowledge concerning its physiology and genetics make Bacillus subtilis an attractive system for strain engineering and analysis. Transposon-based mutagenesis strategies generate large libraries of mutant strains that can be used to investigate molecular mecha ...

Screening for large gene rearrangements has become established as an important part of molecular medicine; however, it is also challenging as these rearrangements range from an extra copy of a complete chromosome(s) to deletion or duplication of a single exon. In this chapter, we describe a ver ...