Maruyama, N. | bioRxiv Science

bioRxiv2024

DNA-free and genotype-independent CRISPR/Cas9 system in soybean

The CRISPR/Cas9 system has revolutionized the field of plant genetic engineering. Here we report a smart genome editing system of soybean by using iPB-RNP method without introducing foreign DNA and requiring traditional tissue culture processes such as embryogenesis and organogenesis. Shoot apical meristem (SAM) of embryonic axes was used the target tissue for genome editing, because the SAM in soybean mature seeds has stem cells and specific cell layer developing germ cells during reproductive growth stage. In the iPB-RNP method, the complex of ribonucleoprotein (RNP) and Cas9 protein was directly delivered into SAM stem cells via particle bombardment and genome-edited plants were generated from these SAMs. Soybean allergenic gene Gly m Bd 30K, which we previously generated genome-editing soybean by using Agrobacterium-mediated transformation and particle bombardment in our previous studies, was targeted in this study. Many E0 (the first generation of genome-edited) plants in this experiment harbored mutant alleles at the targeted locus. Editing frequency of inducing mutations transmissible to the E1 generation was approximately 0.4 to 4.6 % of all E0 plants utilized in various soybean varieties. Furthermore, Gly m Bd 30K protein in mature seeds was not detected by western blot analysis due to flame-sift mutations. Our results offer a practical approach for both plant regeneration- and DNA-free genome editing achieved by delivering RNP into the SAM of dicotyledonous plants.

plant biology↗

bioRxiv2020

The transcriptomic landscapes of diverse rice cultivars grown under mild drought conditions

Root system architecture affects plant drought tolerance and other key agronomic traits such as lodging. However, although phenotypic and genomic variation has been extensively analyzed, few field studies have integrated phenotypic and transcriptomic information, especially for below-ground traits such as root system architecture. Here, we report the phenotypic and transcriptomic landscape of 61 rice (Oryza sativa) accessions with highly diverse below-ground traits grown in an upland field under mild drought stress. We found that four principal components explained the phenotypic variation and that accessions could be classified into four admixture groups (admixed, aus, indica, and japonica) based on their tiller numbers and crown root diameters. Transcriptome analysis revealed that differentially expressed genes associated with specific admixture groups were enriched with stress response-related genes, suggesting that admixture groups have distinct stress response mechanisms. Root growth was negatively correlated with auxin-inducible genes, suggesting an association between auxin signaling and mild drought stress. A negative correlation between crown root diameter and stress response-related genes suggested that thicker crown root diameter is associated with resistance to mild drought stress. Finally, co-expression network analysis implemented with DNA affinity purification followed by sequencing (DAP-seq) analysis identified phytohormone signaling networks and key transcription factors negatively regulating crown root diameter. Our datasets provide a useful resource for understanding the genomic and transcriptomic basis of phenotypic variation under mild drought stress. ONE-SENTENCE SUMMARYCatalog of the phenomes and transcriptomes of rice cultivars grown in upland fields provides a resource for further studies toward breeding climate-resilient crops.

plant biology↗