Zan Lingxiong, Liu Haidong, Zhao Xutao, Du Dezhi, Li Kaixiang
Academy of Agricultural and Forestry Sciences of Qinghai University, Xining 810016, China.
Laboratory for Research and Utilization of Qinghai Tibet Plateau Germplasm Resources, Xining 810016, China.
Curr Issues Mol Biol. 2024 Dec 30;47(1):15. doi: 10.3390/cimb47010015.
The determinate inflorescence trait of L. is associated with various desirable agricultural characteristics. ( and ), which encode the transcription factor have previously been identified as candidate genes controlling this trait through map-based cloning. However, the mechanism underlying the effects of the BnTFL1 proteins remains unclear. Further, proteins generally interact with each other to fulfill their biological functions. The objective of this study was to construct a cDNA library of the shoot apical meristem (SAM) of . and screen for proteins that interact with BnTFL1s, to better understand its mechanism of action. The recombination efficiency of the yeast two-hybrid (Y2H) library that we constructed was 100%, with insertion fragment lengths ranging from 750 to 2000 bp and a capacity of approximately 1.44 × 10 CFUs (colony-forming units), sufficient for screening protein interactions. Additionally, the bait vector pGBKT7-BnTFL1s was transformed into yeast cells alongside positive and negative controls, demonstrating no toxicity to the yeast cells and no self-activation. This bait was used to screen the SAM cDNA library of , ultimately identifying two BnTFL1s-interacting proteins: 14-3-3-like protein GF14 omega GRF2. These interactions were verified through one-to-one interaction experiments. This study provides a foundation for further research on the biological functions of the genes and their regulatory role in inflorescence formation in , while providing a reference for studying similar mechanisms in other plants.
L. 的有限花序性状与多种理想的农艺性状相关。（和），它们编码转录因子，先前已通过图位克隆被鉴定为控制该性状的候选基因。然而，BnTFL1 蛋白作用的潜在机制仍不清楚。此外，蛋白质通常相互作用以实现其生物学功能。本研究的目的是构建 L. 茎尖分生组织（SAM）的 cDNA 文库，并筛选与 BnTFL1 相互作用的蛋白质，以更好地了解其作用机制。我们构建的酵母双杂交（Y2H）文库的重组效率为 100%，插入片段长度在 750 至 2000 bp 之间，容量约为 1.44×10 CFU（菌落形成单位），足以筛选蛋白质相互作用。此外，将诱饵载体 pGBKT7-BnTFL1 与阳性和阴性对照一起转化到酵母细胞中，证明对酵母细胞无毒性且无自激活。该诱饵用于筛选 L. 的 SAM cDNA 文库，最终鉴定出两种与 BnTFL1 相互作用的蛋白质：14-3-3 样蛋白 GF14 omega GRF2。这些相互作用通过一对一相互作用实验得到验证。本研究为进一步研究 L. 中相关基因的生物学功能及其在花序形成中的调控作用奠定了基础，同时为研究其他植物中的类似机制提供了参考。
