Modifying multipotent, self-renewing human stem cells with mammalian artificial chromosomes (MACs), present a promising clinical strategy for numerous diseases, especially ex vivo cell therapies that can benefit from constitutive or overexpression of therapeutic gene(s). ...

Current transgenic technologies for gene transfer into the germline of mammals cause a random integration of exogenous naked DNA into the host genome that can generate undesirable position effects as well as insertional mutations. The vectors used to generate transgenic animals are l ...

Horizontal gene transfer or simply transgenic technology has evolved much since 1980. Gene delivery strategies, systems, and equipments have become more and more precise and efficient. It has also been shown that even chromosomes can be used besides traditional plasmid and viral vectors ...

Gene therapy encounters important problems such as insertional mutagenesis caused by the integration of viral vectors. These problems could be circumvented by the use of mammalian artificial chromosomes (MACs) that are unique and high capacity gene delivery tools. MACs were delive ...

Mammalian satellite DNA-based artificial chromosomes (SATACs) are unique among the mammalian artificial chromosomes. These reproducibly generated de novo chromosomes are stably maintained in different species, readily purified from the host cell’s chromosomes and can be i ...

Among the many techniques of cloning new genes, one approach involves degenerate primers (1–7). The approach usually requires the following three steps: 1. Using degenerate primers to amplify part of the gene of interest by PCR: The degenerate pri

A crucial and often decisive test of a nuclear gene being involved in a given process is the complementation of mutants. Restoring the wild type phenotype by the wild type gene introduced into the mutant is a major piece of evidence for the function of this gene. We have developed a rapid and reliable method to ...

Although type I topoisomerases do not require a high-energy cofactor, type II topoisomerases require ATP in order to carry out their essential catalytic functions (1–4). ATP binding is necessary to close the protein clamp (5,6) and trigger DNA strand passage (7,8), whereas hydrolysis is neces ...

This chapter outlines the generation and application of human papillomavirus type 33 (HPV33) pseudovirions. The method describes (1) the construction of vaccinia viruses recombinant for the major and minor HPV capsid proteins, L1 and L2, respectively; (2) the transfection of Cos7 cells ...

The extraordinary sensitivity of reverse-transcriptase-polymerase chain reaction (RT-PCR) makes it a powerful technique for specific mRNA detection, particularly when tissue availability is limiting, and when the mRNA to be detected is present in low abundance. A disadvantage ...

The completion of whole-genome sequencing projects offers the opportunity of creating high-resolution maps of specific segments in a known genomic DNA sequence. For this purpose, several genome browsers have been created. They include the map-view (http:// www.ncbi.nlm.nih.gov/m ...

At any particular point in time, the full complement of transcribed RNAs and relevant proteins of a cell are known as the transcriptome and proteome, respectively. The composition of these two populations changes throughout the life cycle of a parasite or in response to environmental factors, ...

DNA microarray platforms represent a functional genomics technology that uses structured information obtained from genomic sequencing efforts as a means to study transcriptional processes in a systematic and high-throughput manner. Specifically in this chapter, we outline ...

The genome sequencing of protozoan parasites has facilitated the development of powerful postgenomics tools such as DNA microarrays and revolutionized the study of parasite biology. Large-scale genomic comparisons are useful in identifying the extent of genomic variability a ...

Genotyping is an important tool for epidemiological and population genetic studies in protozoan parasites. The most commonly used method for genotyping is polymerase chain reaction (PCR)-based restriction fragment length polymorphism (RFLP) analysis of single nucleotide p ...

Most members of the AID/APOBEC family of polynucleotide deaminases can catalyse the deamination of cytosine to uracil in DNA. They thereby function as active DNA mutators. Here, we describe how bacterial papillation assays can be adapted to monitor the mutator activity of AID/APOBEC prot ...

Human APOBEC3G (A3G) is a cytidine deaminase that broadly restricts the replication of many retroviruses, including HIV-1. In different cell types, cytoplasmic A3G is expressed in high-molecular-mass (HMM) RNA–protein complexes or low-molecular-mass (LMM) forms displaying dif ...

RNA editing deaminases act on a variety of targets in different organisms. A number of such enzymes have been shown to act on mRNA, with the resultant nucleotide changes modifying a transcript’s information content. Though the deaminase activity of mRNA editing enzymes is readily demonstra ...

Substitutional RNA editing represents an important posttranscriptional enzymatic pathway for increasing genetic plasticity by permitting production of different translation products from a single genomically encoded template. One of the best-characterized examp ...

mRNA editing in plastids (chloroplasts) of higher plants proceeds by cytidine-to-uridine conversion at highly specific sites. Editing sites are recognized by the interplay of cis-acting elements at the RNA level and site-specific trans-acting protein factors that are believed to b ...