DescriptionThe retrotransposons – the mobile genetic elements comprise the bulk of all genomes of eukaryotes. They generally show widespread chromosomal dispersion, variable copy number and random distribution in the genome. Retrotransposons move to new chromo-somal locations via an RNA intermediate, and insert new cDNA copies back into the genome. In higher plants, more than half of the repetitive DNA consists of retrotrans-posons, a component dynamic by its ability to integrate new copies and facilitate to ho-mologous recombination. They dynamic and playing role in chromosome crossing over recognition and in recombination DNA between homologous chromosomes. Different retrotransposon families, each with its own lineage and structure, have the potential to have been active at distinct phases in the evolution of a species. The sequences of re-trotransposons carry the promoters, which bind the nuclear factors of initialization of a transcription and initiated the RNA synthesis by polymerases II and III. The most part of retrotransposon sequences are inactivated by mutations and partially transcribed. The different variants of retrotransposons can be completely inactive, seldom or constantly active. In natural populations of plants activity of the majority retrotransposon much more above, than at cultivated, domestic forms. High polymorphism at the natural popu-lations, revealed by PCR use based on conservative retrotransposons sequences are hid-den in phenotype. Retrotransposons can be used for markers because integration of a daughter copy creates new joints between genomic DNA and the conserved LTRs. Var-ious molecular marker systems have been developed that exploit the ubiquitous nature of these genetic elements and their property of stable integration into dispersed chromoso-mal loci that are polymorphic within species. To detect polymorphisms for retrotrans-poson insertions, marker systems generally rely on PCR amplification between the re-trotransposon termini and some component of flanking genomic DNA. The main meth-ods of IRAP, REMAP, RBIP, and SSAP all detect the polymorphic sites at which the retrotransposon DNA is integrated into the genome. Marker systems exploiting these methods can be easily developed and are inexpensively deployed in the absence of ex-tensive genome sequence data. Here, we describe protocols for the IRAP, REMAP and iPBS techniques, including methods for PCR amplification with a single primer or with two primers, agarose gel electrophoresis of the product using optimal electrophoresis buffers, we also describe iPBS techniques for the rapid isolation of retrotransposon ter-mini and full-length elements.
|Aug 13 2020
|Gamov conference in Odessa National University
|Degree of Recognition