In order to restore or to introduce a gene function integrating viral vector systems are used to genetically modify hematopoietic stem cells. The occurrence of immortalized cell clones after transduction in vitro (Blood 106:3932–3939, 2005) and clonal dominance as well as leukemia in pre ...
Identification of retroviral vector insertion sites in single, dominating cell clones has become an important tool for the investigation of cellular signalling pathways involved in clonal expansion and malignant transformation. Also, recent severe adverse events in clinical ...
Identification, monitoring, and analysis of genetically modified cells in the peripheral blood are an important component of the clinical follow-up of patients treated by hematopoietic cell gene therapy. Analysis of gene-marked peripheral blood cells provides crucial inform ...
In recent years, one of the most quickly incorporated methods in biomedical research has been microarray technology. Microarrays have been designed for the purpose of genotyping (e.g., SNP analysis), expression analysis (mRNA, miRNA, exon arrays), chromatin immunoprecipitations ...
Maintaining the stability of the genome is critical to cell survival and normal cell growth. Genetic modification of hematopoietic cells might bear an inherent increased risk for the accumulation of DNA mutations. It frequently requires cultivation of the cells under super-physiol ...
Complex biological samples hold significant information on the health status and on development of disease. Approximately 35,000 human genes give rise to more than 1,000,000 functional entities at the protein level. Thus, the proteome provides a much richer source of information than the ...
Hematopoietic stem cells (HSC) are rare with estimated frequencies of 1 in 10,000 bone marrow cells and 1 in every 100,000 blood cells. The most important characteristic of HSC is their capacity to provide complete restoration of all blood cell lineages after bone marrow ablation. Therefore th ...
Cytogenetic characterization of murine chromosomes using banding techniques like R- or G-banding is technically demanding due to the similar size and the acrocentric structure of all chromosomes. The molecular cytogenetic technique of spectral karyotyping (SKY) overcomes th ...
The nonhuman primate is a highly relevant model for the study of human diseases, and currently there is a significant need for populations of animals with specific genotypes that can not be satisfied by the capture of animals from the wild or by conventional breeding. There is an even greater need for ge ...
Cloning using somatic cell nuclear transfer (SCNT) may be a useful tool for conserving genetic diversity and for propagating exotic and/or endangered animal species. Somatic cells can be obtained easily, expanded in culture, cryopreserved, and thawed at a later date for use in NT. Significa ...
Apart from the biological and ethical problems, technical difficulties also hamper the improvement and widespread application of somatic cell nuclear transfer (NT). Recently introduced zona-free procedures may offer a solution for the latter problem. The most radical approach of ...
For many years the lack of germline competent embryonic stem cell lines in livestock meant that the targeted modification of endogenous genes was not possible in these species. The demonstration that livestock could be cloned by nuclear transfer from cultured somatic cells has now provid ...
Nuclear transfer (NT) using transgenic donor cells is an efficient means for generation of transgenic founder goats, especially in regard to the number of animals required to produce a transgenic founder expressing the protein of interest. Vectors can be designed for organ-specific exp ...
When immature bovine oocytes are released from their follicles and are cultured in standard maturation medium, they resume the first meiotic division. The alteration of basic maturation conditions can affect oocyte competence significantly, as reflected by the morula and blastoc ...
This chapter traces the orgin and progress of nuclear transfer that later became the paradigm for cloning animals. Classic studies in cytology, embryology, or genetics spanning more than five centuries that led to nuclear transfers in unicellular animals and to those in oocytes of insects, f ...
Somatic cloning technology involves the transfer of a somatic cell nucleus into an enucleated oocyte, followed by activation and in vitro culture. Efficiency in terms of live offspring generally remains very low. Little attention has been devoted so far to the impact of culture environment ...
After nuclear transfer, the recipient oocyte must be stimulated to initiate development. This stimulation is achieved by inducing changes in the oocyte cytoplasm that normally are triggered by the sperm during fertilization. In most cases, such changes include a transient increase in t ...
Interest is increasing in assisted reproductive technologies in the pig involving embryo cryopreservation, cloning, and genetic modification. Although inherently inefficient and variable in their outcome, the successful application of these techniques in this species is c ...
The management of clone-bearing recipients and neonatal clones is a critical component of successful cloning of mammals by nuclear transfer. The methodology discussed in this chapter is based largely on a double corticosteroid regime to induce parturition and fetal organ maturation ...
The stage of the donor cell cycle is a major factor in the success of cloning. Quiescent cells arrested in the G0/G1 phases of the cell cycle by either serum starvation or growth arrest when cultured cells reach confluence have been used as donors to produce cloned animals. Recently, we have developed a nov ...
