[DNA genotyping of oral epithelial cells by laser capture microdissection].
ABSTRACT The STR genotypping of trace oral epithelial cells which are microdissected by laser capture microdissection system (LCM) is explored.
The oral epithelial cells are microdissected using a low-power infrared laser by VERITAS Microdissection Instrument. STR loci of Profiler Plus are detected by multiplex PCR procesures.
DNA genotyping of 7-8 oral epithelial cells are succeeded, and DNA genotyping of 3-4 oral epithelial cells are failed.
It is viable in genotyping of trace oral epithelial cells by Laser Capture Microdissection as a new technology of seperating single cell.
- SourceAvailable from: Igor V Sharakhov[Show abstract] [Hide abstract]
ABSTRACT: Fluorescent in situ hybridization (FISH) of whole arm chromosome probes is a robust technique for mapping genomic regions of interest, detecting chromosomal rearrangements, and studying three-dimensional (3D) organization of chromosomes in the cell nucleus. The advent of laser capture microdissection (LCM) and whole genome amplification (WGA) allows obtaining large quantities of DNA from single cells. The increased sensitivity of WGA kits prompted us to develop chromosome paints and to use them for exploring chromosome organization and evolution in non-model organisms. Here, we present a simple method for isolating and amplifying the euchromatic segments of single polytene chromosome arms from ovarian nurse cells of the African malaria mosquito Anopheles gambiae. This procedure provides an efficient platform for obtaining chromosome paints, while reducing the overall risk of introducing foreign DNA to the sample. The use of WGA allows for several rounds of re-amplification, resulting in high quantities of DNA that can be utilized for multiple experiments, including 2D and 3D FISH. We demonstrated that the developed chromosome paints can be successfully used to establish the correspondence between euchromatic portions of polytene and mitotic chromosome arms in An. gambiae. Overall, the union of LCM and single-chromosome WGA provides an efficient tool for creating significant amounts of target DNA for future cytogenetic and genomic studies.Journal of Visualized Experiments 01/2014; DOI:10.3791/51173