National Key Laboratory of Crop Genetic Improvement, Hubei Hongshan Laboratory, Huazhong Agricultural University, Wuhan, Hubei, 430070, China.
Key Laboratory of Oasis Ecology Agricultural of Xinjiang Production and Construction Corps, Agricultural College, Shihezi University, Shihezi, Xinjiang, 832003, China.
Genome Biol. 2024 Feb 26;25(1):59. doi: 10.1186/s13059-024-03189-8.
CRISPR/Cas-derived base editor enables precise editing of target sites and has been widely used for basic research and crop genetic improvement. However, the editing efficiency of base editors at different targets varies greatly.
Here, we develop a set of highly efficient base editors in cotton plants. GhABE8e, which is fused to conventional nCas9, exhibits 99.9% editing efficiency, compared to GhABE7.10 with 64.9%, and no off-target editing is detected. We further replace nCas9 with dCpf1, which recognizes TTTV PAM sequences, to broaden the range of the target site. To explore the functional divergence of TERMINAL FLOWER 1 (TFL1), we edit the non-coding and coding regions of GhTFL1 with 26 targets to generate a comprehensive allelic population including 300 independent lines in cotton. This allows hidden pleiotropic roles for GhTFL1 to be revealed and allows us to rapidly achieve directed domestication of cotton and create ideotype germplasm with moderate height, shortened fruiting branches, compact plant, and early-flowering. Further, by exploring the molecular mechanism of the GhTFL1 and GhTFL1 mutations, we find that the GhTFL1 mutation weakens the binding strength of the GhTFL1 to other proteins but does not lead to a complete loss of GhTFL1 function.
This strategy provides an important technical platform and genetic information for the study and creation of ideal plant architecture.
CRISPR/Cas 衍生的碱基编辑器可实现靶位点的精确编辑，已广泛用于基础研究和作物遗传改良。然而，碱基编辑器在不同靶标上的编辑效率差异很大。
本研究在棉花中开发了一套高效的碱基编辑器。与 GhABE7.10 的 64.9%相比，融合常规 nCas9 的 GhABE8e 的编辑效率达到 99.9%，并且未检测到脱靶编辑。我们进一步用识别 TTTV PAM 序列的 dCpf1 替代 nCas9，以拓宽靶位点范围。为了探索 TERMINAL FLOWER 1（TFL1）的功能分化，我们对 GhTFL1 的非编码区和编码区进行了 26 个靶点的编辑，生成了一个包含 300 个独立系的 GhTFL1 综合等位基因群体。这揭示了 GhTFL1 的隐藏多效性作用，使我们能够快速实现棉花的定向驯化，并创建具有适度高度、缩短果枝、紧凑植株和早花的理想型种质资源。此外，通过探索 GhTFL1 和 GhTFL1 突变的分子机制，我们发现 GhTFL1 突变削弱了 GhTFL1 与其他蛋白质的结合强度，但不会导致 GhTFL1 功能完全丧失。
该策略为理想植物结构的研究和创造提供了重要的技术平台和遗传信息。
