Michael Smith Laboratories, University of British Columbia, Vancouver, BC, V6T 1Z4, Canada.
Department of Botany, University of British Columbia, Vancouver, BC, V6T 1Z4, Canada.
Plant J. 2024 Jan;117(1):161-176. doi: 10.1111/tpj.16482. Epub 2023 Sep 29.
Agrobacterium-mediated transformation enables random transfer-DNA (T-DNA) insertion into plant genomes. T-DNA insertion into a gene's exons, introns or untranscribed regions close to the start or stop codon can disrupt gene function. Such T-DNA mutants have been useful for reverse genetics analysis, especially in Arabidopsis thaliana. As T-DNAs are inserted into genomic DNA, they are generally believed to be stably inherited. Here, we report a phenomenon of reversion of intronic T-DNA mutant phenotypes. From a suppressor screen using intronic T-DNA pi4kβ1,2 double mutant, we recovered intragenic mutants of pi4kβ1, which suppressed the autoimmunity of the double mutant. These mutants carried deletions in the intronic T-DNAs, resulting in elevated transcription of normal PI4Kβ1. Such reversion of T-DNA insertional mutant phenotype stresses the need for caution when using intronic T-DNA mutants and reiterates the importance of using irreversible null mutant alleles in genetic analyses.
农杆菌介导的转化能够实现随机转移 DNA（T-DNA）插入植物基因组。T-DNA 插入到基因的外显子、内含子或靠近起始或终止密码子的非转录区域，可以破坏基因功能。这种 T-DNA 突变体在反向遗传学分析中非常有用，尤其是在拟南芥中。由于 T-DNAs 插入到基因组 DNA 中，它们通常被认为是稳定遗传的。在这里，我们报告了内含子 T-DNA 突变体表型回复的现象。从使用内含子 T-DNA pi4kβ1,2 双突变体的抑制子筛选中，我们恢复了 pi4kβ1 的基因内突变体，该突变体抑制了双突变体的自身免疫。这些突变体在内含子 T-DNAs 中携带缺失，导致正常 PI4Kβ1 的转录升高。这种 T-DNA 插入突变体表型的回复强调了在使用内含子 T-DNA 突变体时需要谨慎，并再次强调了在遗传分析中使用不可逆的 null 突变等位基因的重要性。
