Know Southern blotting
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The Southern blot is a technique that uses gel electrophoresis combined with labeled probes for a DNA sequence of interest that allows for the detection of repeat expansions within specific genes.
Southern blotting has application in instances where the structure of a gene (cloned or uncloned) is to be investigated. It may be used to determine a restriction enzyme map of a gene or to investigate natural variation or mutational events in a gene such as insertions deletions or rearrangements resulting from recombination. If appropriate controls are included and the procedure is carried out with care, Southern blotting may be used in a quantitative fashion to investigate phenomena such as gene amplification or loss of heterozygosity. Dimorphism of restriction enzyme sites within the population which results in restriction fragment length polymorphisms (RFLPs) was first discovered by Southern blotting and this later led to the discovery of other forms of variation in genomes that have been crucial to the mapping of genes and to our understanding of mutational processes.
With regard to immunology, the principal application of Southern blotting remains the investigation of the clonal rearrangements of the immunoglobulin and T cell receptor genes involved in the immune response. This can be illustrated by considering B lymphocytes, each of which carry a productively rearranged immunoglobulin allele as well as an allele with the germline structure or one that has undergone a nonproductive rearrangement. Each rearrangement event will be specific to a single cell in the population of B cells and Southern blot analysis of total B cell DNA would reveal only the germline arrangement of immunoglobulin genes as any individual rearranged alleles would be represented negligibly. As a result, bands from rearranged alleles fall far below detection limits. The structure of rearranged genes only becomes apparent by Southern blotting where there has been clonal expansion of a single progenitor B cell such as occurs in lymphoproliferative disorders. In such instances, Southern blotting may be used to characterize the structure of the rearranged gene in the expanded clone.