ABSTRACT
RNA directed fluorescence in situ hybridization (FISH) is an effective approach to localize RNA sequences in situ, visualize gene transcripts, and estimate gene expression in time and space. In a typical interphase nucleus, transcribed genes, together with nascent transcripts at the site of their synthesis, are visible as dot-like foci., in fishes, amphibians, reptiles, birds, and certain insects, hypertranscription during oogenesis leads to decompactization of hundreds of transcribed genomic regions and transformation of the meiotic chromosomes into highly elongated lampbrush chromosomes (LBCs). This chapter provides an effective RNA-FISH protocol that includes steps essential to preserve RNA on LBC preparations; also presents necessary control experiments to estimate gene expression. The methodology enables simultaneous visualization of up to 10 transcribed single-copy genomic loci on the same LBC. Conventional 2D fluorescence microscopy is used to examine LBC preparations after FISH.
