Ds qi2 microscopy camera

Young anthers during meiotic stages were fixed in 4% (w/v) paraformaldehyde for 30 min at room temperature and stored in 1x Buffer A at 4°C. Immunofluorescence was performed as previously described (Pawlowski et al., 2003 (link); Cheng, 2013 (link)). The primary antibodies against ASY1, ZYP1, and γH2AX were prepared as described previously (Jing et al., 2019 (link)). Antibody against RAD51 was a gift from Wojtek Pawlowski’s Lab at Cornell University. Fluorochrome-coupled secondary antibodies (ABclonal) were used for fluorescence detection. All primary and secondary antibodies were diluted at 1:100. Images of meiocytes were observed and captured using a Ci-S-FL microscope (Nikon) equipped with a DS-Qi2 microscopy camera (Nikon, Tokyo, Japan). The images were captured by software NIS-Elements and colored by the ImageJ software.

The FISH analysis was performed according to protocols described previously (Richards and Ausube, 1988 (link); Li and Arumuganathan, 2001 (link); Wang et al., 2006a (link); Han et al., 2007 (link); Cheng, 2013 (link)6a). Two repetitive DNA elements, 5S rDNA repeats (pTa794) and the telomere-specific repeats (pAtT4), were used as probes (Richards and Ausube, 1988 (link)). Probes were labeled with digoxigenin by nick translation mix (Roche) and detected with anti-digoxigenin antibody (Vector). Chromosome images were captured under a Ci-S-FL fluorescence microscope (Nikon) equipped with a DS-Qi2 microscopy camera (Nikon, Tokyo, Japan).