2D and 3D Chromosome Painting in Malaria Mosquitoes
Sharakhov, Igor V.
MetadataShow full item record
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.
Showing items related by title, author, creator and subject.
Identification and Characterization of Y Chromosome and M Locus Genes in Anopheles and Aedes Mosquitoes Using the Chromosome Quotient Method Hall, Andrew Brantley (Virginia Tech, 2016-03-22)In mosquitoes, sex determination is initiated by a dominant male-determining factor located on the Y chromosome in Anopheles mosquitoes or in a small Y-like region called the M locus in Aedes mosquitoes. Before my research, ...
Mitotic Dynamics of Normally and Mis-attached Chromosomes and Post-mitotic Behavior of Missegregated Chromosomes He, Bin (Virginia Tech, 2015-06-01)Equal segregation of the replicated genomic content to the two daughter cells is the major task of mitotic cells. The segregation is controlled by a complex system in the cell and relies mainly on the interaction between ...
Kinney, Nicholas A.; Sharakhov, Igor V.; Onufriev, Alexey V. (2018-01-22)Abstract Background It is well recognized that the interphase chromatin of higher eukaryotes folds into non-random configurations forming territories within the nucleus. ...