Retroviral Insertion Site Analysis in Dominant Haematopoietic Clones
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 trials resulting from retroviral vector-mediated insertional mutagenesis underline the need of well-designed safety studies including integration site analyses to estimate cost/benefit ratios in gene therapy. We have recently described a modified ligation-mediated PCR (LM PCR) method allowing preferential retrieval of insertion sites causally linked to clonal dominance of an affected clone. In the first part of the given work we focus on particularities of the LM PCR procedure to be taken into account when working with self-inactivating as compared to ‘classical’ retrovectors. In the following sections we focus on data acquisition, processing, organisation, and analysis. Thus the protocol presented here should be helpful in establishing and utilising databases of retroviral integration sites.
- Fluorescence Procedures for the Actin and Tubulin Cytoskeleton in Fixed Cells
- 常用细胞凋亡检测方法(二)
- Tools for Predicting Binding and Insertion of CPPs into Lipid Bilayers
- Analysis of Molecular Chaperone Activities Using In Vitro and In Vivo Approaches
- Fertilization and Sperm Chemotaxis in Ascidians
- 细胞培养中常见的污染情况
- Random Mutagenesis by Using Mixtures of dNTP and dITP in PCR
- Retroviral Insertion Site Analysis in Dominant Haematopoietic Clones
- Preparation of Astrocyte, Oligodendrocyte, and Microglia Cultures from Primary Rat Cerebral Cultures
- Monitoring Oncogenic B-RAF-Induced Senescence in Melanocytes