Using RNA FISH to study gene expression during mammalian meiosis.
During mouse meiosis, gene expression and homologous synapsis are intimately linked. Chromosomes that fail to synapse at the zygotene-pachytene transition become transcriptionally silenced by a process called Meiotic Silencing of Unsynapsed Chromatin (MSUC), and this silencing (or defects in it) may in turn cause germ cell losses and infertility. Gene transcription at the chromosomal level can be readily observed using RNA fluorescence in-situ hybridisation (FISH), and this approach allows one to directly compare expression at a specific locus with the synaptic status of the chromosome domain on which it resides. Here we describe a protocol for carrying out RNA FISH on male meiotic cells, together with detail on the important controls and common problems associated with this technique.
Mahadevaiah
Shantha K
SK
Division of Stem Cell Biology and Developmental Genetics, MRC National Institute for Medical Research, London, UK.
Costa
Yael
Y
Turner
James M A
JM
D000818:Animals; D022202:Chromosomes, Artificial, Bacterial / Q000737:chemistry / Q000502:physiology; D015870:Gene Expression / Q000502:physiology; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D008297:Male; D008322:Mammals / Q000235:genetics*; D008540:Meiosis / Q000235:genetics* / Q000502:physiology; D051379:Mice; D015347:RNA Probes / Q000032:analysis / Q000502:physiology*
D015347:RNA Probes