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Abstract Key messageBiolistic particle bombardment was used to deliver CRISPR-Cas9 ribonucleoprotein complexes (RNP) into the shoot apical meristem tissue of citrus and axillary meristem tissue of poplar, generating directed mutations in target genes. AbstractThe use of meristematic tissues offers a strategic approach to genome editing in woody species, especially those that are recalcitrant to conventional tissue culture, as these regions contain totipotent, highly regenerative cells capable of giving rise to whole plants. Here, we employed biolistic delivery of genome-editing reagents into theshoot apical meristem (SAM) of citrus and the axillary meristems (AXM) of poplar. The system was first validated using a GFP expression construct and subsequently applied for targeted genome editing. In citrus, edited plants were obtained at the CsNPR3 locus exclusively through the delivery of CRISPR/Cas9 ribonucleoproteins (RNPs), whereas plasmid-based vectors were unsuccessful. Similarly, genome editing in poplar was achieved using RNPs targeting the Pt4CL1 gene. Although chimeric events were detected, this approach provides a feasible and innovative framework for producing transgene-free edited perennial plants. 

Although climate can strongly influence erosional efficiency (i.e., erosion rate for a given topography), demonstrating its impact in tectonically active areas has been challenging due to other confounding controlling factors, such as lithology. Here, we show that 10Be-derived erosion rates and efficiencies in the Himalayan orogen exhibit distinct relationships with climatic factors depending on lithology. We compile 173 10Be-derived, basin- averaged erosion rates across the orogen, including 12 newly measured rates from the Dibang and Lohit valleys in the easternmost Himalaya, regions characterized by high precipitation magnitudes and variability. We group basins based on lithologies separated by orogen-scale thrust faults and quantify erosional efficiency coefficients based on the relationships between erosion rates and topographic metrics. Our results show that erosion rates and erosional efficiency from sedimentary and metasedimentary rocks along the Himalayan range front display a positive, nonlinear correlation with climatic factors, such as the number of extreme rainfall events and mean annual precipitation rates. In contrast, erosion rates from crystalline lithologies in the hanging wall of the Main Central thrust show a strong correlation with fluvial topography, whereas erosional efficiency shows no statistically significant correlation with climatic factors. Rapid erosion rates and high erosional efficiencies in the eastern Himalayan range front are likely driven by extreme precipitation on tectonically active, steep slopes composed of mechanically weak metasedimentary rocks. Our findings highlight the importance of the interplay between controlling factors, which include tectonics, lithology, and climate, that drive surface erosion and influence the topographic evolution of orogenic systems. 

Abstract Key messageTransgene-free genome editing of the gene of interest in citrus and poplar has been achieved by co-editing theALSgene via transient transgene expression of an efficient cytosine base editor. AbstractCRISPR-Cas genome editing systems have been widely used in plants. However, such genome-edited plants are nearly always transgenic in the first generation whenAgrobacterium-mediated transformation is used. Transgene-free genome-edited plants are valuable for genetic analysis and breeding as well as simplifying regulatory approval. It can be challenging to generate transgene-free genome-edited plants in vegetatively propagated or perennial plants. To advance transgene-free genome editing in citrus and poplar, we investigated a co-editing strategy using an efficient cytosine base editor (CBE) to edit theALSgene to confer herbicide resistance combined with transient transgene expression and potential mobile RNA-based movement of CBE transcripts to neighboring, non-transgenic cells. An FCY-UPP based cytotoxin system was used to select non-transgenic plants that survive after culturing on 5-FC containing medium. While the editing efficiency is higher in poplar than in citrus, our results show that the CBE-based co-editing strategy works in both citrus and poplar, albeit with low efficiency for biallelic edits. Unexpectedly, the addition of the TLS mobile RNA sequence reduced genome editing efficiency in both transgenic and non-transgenic plants. Although a small fraction of escaping plants is detected in both positive and negative selection processes, our data demonstrate a promising approach for generating transgene-free base-edited plants.