Figure 3

Masked mRNA is stored within the microspore of M. vestita. a, Microspore fixed after 30 minutes of development, double-stained with DAPI (blue) and PY (green). b, Microspore fixed after 45 minutes of development (just prior to the prothallial division) double-stained with DAPI (blue) PY (green). c, Microspore fixed after 1.5 hours of development double- stained with DAPI (blue) PY (green). d, Microspore fixed after 5 hours of development double-stained with DAPI (blue) PY (green). e-g, Distribution of Poly(A)+ RNA during spermatogenesis. A biotinylated poly(T) probe (red) was used for FISH on microspores fixed at (e) 30 minutes, (f) 1 hour, and (g) 4 hours of development. FISH probes were detected using avidin bound TexasRed. h-m, Microspores fixed after 30 minutes of development. h, FISH using 25mer biotinylated centrin specific probes, and detected using avidin bound Fluorescein (green). i, FISH using 25mer biotinylated SPDS specific probes, and detected using avidin bound Fluorescein (green). j, FISH using traditional biotinylated centrin specific probes, and detected using avidin bound TexasRed (red). k, FISH using traditional biotinylated SPDS specific probes, and detected using avidin bound TexasRed (red). l-m, Microspores pretreated with (l) DNase or (m) RNase and incubated with 25mer biotinylated SPDS probes, which were detected using avidin bound Fluorescein (green). Bar = 25 μm.