Author
Listed:
- Amichai Berman
(Tel Aviv University)
- Ning Su
(University of Chinese Academy of Sciences)
- Zhuorong Li
(University of Chinese Academy of Sciences)
- Udi Landau
(Tel Aviv University)
- Joydeep Chakraborty
(Tel Aviv University)
- Natali Gerbi
(Tel Aviv University)
- Jia Liu
(University of Chinese Academy of Sciences)
- Yuntai Qin
(University of Chinese Academy of Sciences)
- Boxi Yuan
(University of Chinese Academy of Sciences)
- Wei Wei
(University of Chinese Academy of Sciences
Chinese Academy of Sciences)
- Osnat Yanai
(Netter Center)
- Itay Mayrose
(Tel Aviv University)
- Yuqin Zhang
(University of Chinese Academy of Sciences)
- Eilon Shani
(Tel Aviv University)
Abstract
Genetic variance is vital for breeding programs and mutant screening, yet traditional mutagenesis methods wrestle with genetic redundancy and a lack of specificity in gene targeting. CRISPR-Cas9 offers precise, site-specific gene editing, but its application in crop improvement has been limited by scalability challenges. In this study, we develop genome-wide multi-targeted CRISPR libraries in tomato, enhancing the scalability of CRISPR gene editing in crops and addressing the challenges of redundancy while maintaining its precision. We design 15,804 unique single guide RNAs (sgRNAs), each targeting multiple genes within the same gene families. These sgRNAs are classified into 10 sub-libraries based on gene function. We generate approximately 1300 independent CRISPR lines and successfully identify mutants with distinct phenotypes related to fruit development, fruit flavor, nutrient uptake, and pathogen response. Additionally, we develop CRISPR-GuideMap, a double-barcode tagging system to enable large-scale sgRNA tracking in generated plants. Our results demonstrate that multi-targeted CRISPR libraries are scalable and effective for large-scale gene editing and offer an approach to overcome gene functional redundancy in basic plant research and crop breeding.
Suggested Citation
Amichai Berman & Ning Su & Zhuorong Li & Udi Landau & Joydeep Chakraborty & Natali Gerbi & Jia Liu & Yuntai Qin & Boxi Yuan & Wei Wei & Osnat Yanai & Itay Mayrose & Yuqin Zhang & Eilon Shani, 2025.
"Construction of multi-targeted CRISPR libraries in tomato to overcome functional redundancy at genome-scale level,"
Nature Communications, Nature, vol. 16(1), pages 1-14, December.
Handle:
RePEc:nat:natcom:v:16:y:2025:i:1:d:10.1038_s41467-025-59280-6
DOI: 10.1038/s41467-025-59280-6
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