By Nedime Serakinci, Steen Kølvraa (auth.), Thomas Liehr (eds.)
This FISH software advisor presents an summary of the rules and the fundamental options of fluorescence in situ hybridization (FISH) and primed in situ hybridization (PRINS), that are effectively used to review many features of genomic habit and adjustments. In 36 chapters, contributed via overseas specialists of their specific box, the these days a number of techniques and purposes of the robust innovations are awarded and particular protocols are given.
Described listed here are tools utilizing quite a few cellphone kinds and tissues in addition to assorted organisms, corresponding to mammalians, bugs, vegetation and microorganisms. Multicolor FISH techniques and distinctive purposes comparable to the characterization marker chromosomes, breakpoints, cryptic aberrations, nuclear structure and epigenetic adjustments, in addition to (array-based) comparative genomic hybridization experiences are provided. total, the means of selection is brought for unmarried mobile research in human genetics, microbiology, animal and plant sciences.
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Extra resources for Fluorescence In Situ Hybridization (FISH) — Application Guide
Sample text
This functional compartmentalization is achieved as cells enter the cell cycle, and appears to be important in maintaining the heritable repression of a subset of genes. Evidence from several studies has suggested that a DNA binding protein might mediate the silencing and recruitment of genes to heterochromatin. In this way, it been demonstrated that chromosome X inactivation is affected by 16 N. Serakinci, S. Kølvraa a large cis-acting RNA molecule termed the “X inactive-specific transcript” (Xist).
The other area in which the specific virtues of the FISH technique have had a substantial impact is cellular organization and distribution. This impact stems from a very basic feature of FISH, namely that it offers information on the states and locations of specific nucleic acid sequences in morphologically intact structures. This offers unique possibilities to study cellular functions and also cell diversity in a tissue, either as cell-to-cell variations in genome aberrations or as cell-to-cell variations in expressed genes.
Further improvements of the technique were accomplished by Senger and colleagues (1990), who employed an inverted microscope (1,250× magnification) that increased the resolution of the technique and enabled regions to be cut very precisely (about one GTG-band resolution). In addition, the use of a rotating plate and dissection in air made it easier to perform microdissection. Following dissection, chromosomal fragments were de-proteinated, the DNA was digested using restriction enzymes and cloned into plasmids, and then DNA amplification was accomplished with Klenow fragment DNA polymerase.