Desjardins Stuart, Kanyuka Kostya, Higgins James D
Department of Genetics and Genome Biology, University of Leicester, Leicester, UK.
Department of Biointeractions and Crop Protection, Rothamsted Research, Harpenden, UK.
Methods Mol Biol. 2020;2061:319-330. doi: 10.1007/978-1-4939-9818-0_22.
Virus-induced gene silencing (VIGS) is a rapid and cost-effective reverse genetic technology that can be used to assess gene function in wheat. This chapter contains a detailed description of how to target wheat meiotic genes by VIGS. The timing of this technique is critical and has been optimized to silence meiotic genes at peak expression, evidenced by silencing of Triticum aestivum disrupted meiotic cDNA1 (TaDMC1). We also describe cytological techniques that have been adapted for the preparation and analysis of meiocytes in wheat, including fluorescent in situ hybridization (FISH) with directly labeled, synthetic oligonucleotide probes, and immunolocalization on spread material.
病毒诱导的基因沉默（VIGS）是一种快速且经济高效的反向遗传学技术，可用于评估小麦中的基因功能。本章详细描述了如何通过VIGS靶向小麦减数分裂基因。该技术的时机至关重要，并且已经过优化，以便在峰值表达时沉默减数分裂基因，普通小麦减数分裂cDNA1（TaDMC1）的沉默证明了这一点。我们还描述了适用于小麦减数分裂细胞制备和分析的细胞学技术，包括使用直接标记的合成寡核苷酸探针进行荧光原位杂交（FISH），以及在铺展材料上进行免疫定位。
