A study led by Dr. Xiaoqin Liu at the Peking University Institute of Advanced Agricultural Sciences has discovered and characterized a native peanut Ubiquitin4 promoter (AhUBQ4) with strong and consistent transcriptional activity. Recognizing the limitations of foreign promoters like CaMV 35S in peanut transformation — such as gene silencing and expression variability — the team sought a native solution to boost genetic engineering efficiency in this vital crop.

 

 

Using transcriptome data and sequence homology, the authors identified AhUBQ4 as a highly expressed Ubiquitin gene. They cloned a 973-bp promoter fragment and tested its ability to drive gene expression using GUS and Ruby reporters. Transient assays in Nicotiana benthamiana and peanut stem discs, as well as stable transformation in Arabidopsis and peanut hairy roots, confirmed that AhUBQ4 promotes robust, uniform gene expression across diverse tissues and conditions.

 

Further, the AhUBQ4 promoter was used to drive Cas9 expression in a CRISPR/Cas9 system targeting peanut HY5-HOMOLOG genes. Compared to the 35S promoter, the AhUBQ4 promoter achieved stronger GFP expression and higher editing efficiency in transgenic hairy roots, with a broader spectrum of mutation types (Fig. 1).

 

This work establishes the AhUBQ4 promoter as a powerful tool for gene overexpression and precise genome editing in peanut, offering new potential for genetic improvement and molecular breeding in legumes.

 

Fig. 1. Characterization of the AhUBQ4 promoter in transient and stable peanut transformation and its application to multiplex gene editing.

 

Dr. Xiaoqin Liu is the corresponding author. Dr. Yuanyuan Cui and research assistant Qianqian Zhang are co-first authors. Research assistant Qingjing Meng and joint-training MS student Xiaoyu Liu also contributed to the project. Funding was provided by the Shandong Provincial Key R&D Program (2024LZGC035), Taishan Scholar Program (tsqn202103161) and Weifang Science & Technology Development Program (2024JZ001).