Advanced microdissection of chromosomes

St. Petersburg State University Press Service

A group of scientists from St. Petersburg University and F. Schiller University (Germany) under the guidance of Associate Professor of St. Petersburg State University Alla Krasikova has developed a unique method of microdissection of chromosomes. With its help, it became possible to obtain and decipher the smallest fragments of genomic DNA. The results of the study are published in the highly rated scientific journal in the field of genome studies BMC Genomics (Zlotina et al., Microdissection of lampbrush chromosomes as an approach for generation of locus-specific FISH-samples and samples for high-throughput sequencing).

Currently, there is a wide range of cytogenetic and cytological approaches to the study of chromosomes, one of which is the method of mechanical microdissection. This method allows you to isolate individual chromosomes or their individual regions using special glass needles. Then the resulting material can be amplified, that is, to create many copies of it, and analyzed using molecular cytogenetic methods or high-throughput sequencing methods (decoding genomic sequences). Microdissection is most often used to identify chromosomes, accurately determine the boundaries of evolutionary and clinically significant chromosomal rearrangements, to study the spatial organization of the cell nucleus, etc.

Typically, highly compact mitotic chromosomes are used for microdissection, however, due to their small size, the standard approach to microdissection does not allow the study of tiny and complexly organized chromosomal regions. Scientists of St. Petersburg State University together with colleagues from the F. Schiller University managed to develop a unique protocol for microdissection of giant transcriptionally active chromosomes. This method makes it possible to obtain highly specific probes for fluorescent in situ hybridization (that is, localization on chromosome preparations), as well as to create samples for subsequent high-performance sequencing and mapping of the smallest DNA fragments in the corresponding genome.

"The method of chromosome microdissection developed by us made it possible to isolate chromosomal regions with a size of only 1.5–4 million nucleotide pairs," explains Anna Zlotina, the first author of the article, a junior researcher at St. Petersburg State University. – At the same time, a microscopic fragment of only one copy of the chromosome was sufficient for successful amplification. Previously, for microdissection of metaphase chromosomes, scientists used at least 10-15 dissected copies of a much larger chromosomal region as the starting material."

This high result was obtained thanks to the successful cooperation of St. Petersburg State University with the Thomas Lear Laboratory within the framework of the current interuniversity exchange program. In the course of the work, the resources of two major scientific centers were involved: equipment for mechanical microdissection of chromosomes of the F. Schiller University and equipment for high-precision cytogenetic analysis and decoding of genome fragments, which is part of the St. Petersburg State University Science Park.

"The new protocol has already found application in the field of studying giant chromosomes such as lamp brushes," says Alla Krasikova, associate professor at St. Petersburg State University. – It allows you to obtain DNA and RNA samples from specific regions of such chromosomes and determine the exact genomic position, as well as the length and gene composition of isolated fragments. We have managed to develop an approach that is unique in terms of technical performance and effectiveness, with the help of which it is possible to correlate certain cellular structures and even individual genes with known DNA sequences of the genome. It can also serve as a convenient tool for a detailed study of functionally significant regions of chromosomes and the mechanisms of realization of genetic information."

Portal "Eternal youth" http://vechnayamolodost.ru  09.03.2016