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Efficient scar-free knock-ins of several kilobases in plants by engineered CRISPR-Cas endonucleases

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Molecular Plant 2024年第5期17卷 824-837页

作者： Tom schreiber Anja Prange Petra Schafer Thomas Iwen Ramona Grutzner Sylvestre Marillonnet Aurelie Lepage Marie Javelle Wyatt paul Alain Tissier Department of Cell and Metabolic Biology Leibniz Institute of Plant BiochemistryWeinberg 306120 Halle(Saale)Germany Limagrain Centre de RechercheRoute d’EnnezatCS 9012663720 ChappesFrance

In plants and mammals,non-homologous end-joining is the dominant pathway to repair DNA doublestrand breaks,making it challenging to generate knock-in *** this study,we identified two groups of exonucleases from the herpes virus and the bacteriophage T7 families that conferred an up to 38-fold increase in homology-directed repair frequencies when fused to Cas9/Cas12a in a tobacco mosaic virus-based transient assay in Nicotiana *** achieved precise and scar-free insertion of several kilobases of DNA both in transient and stable transformation *** Arabidopsis thaliana,fusion of Cas9 to a herpes virus family exonuclease led to 10-fold higher frequencies of knock-ins in the first generation of *** addition,we demonstrated stable and heritable knock-ins in wheat in 1%of the primary *** together,our results open perspectives for the routine production of heritable knock-in and gene replacement events in plants.

关键词： homology-directed repair knockin gene replacement CRISPR-Cas 50 exonuclease plants

Apoplastic barriers of Populus×canescens roots in reaction to different cultivation conditions and abiotic stress treatments

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Stress Biology 2023年第1期3卷 269-282页

作者： paul Grunhofer Ines Heimerich Lena Herzig Svenja Pohl Lukas schreiber Department of Ecophysiology Institute of Cellular and Molecular BotanyUniversity of BonnKirschallee 153115 BonnGermany

Populus is an important tree genus frequently cultivated for economical ***,the high sensitivity of poplars towards water deficit,drought,and salt accumulation significantly affects plant productivity and limits biomass *** cultivation and abiotic stress conditions have been described to significantly induce the formation of apoplastic barriers(Casparian bands and suberin lamellae)in roots of different monocotyledonous crop ***,this study aimed to investigate to which degree the roots of the dicotyledonous gray poplar(Populus×canescens)react to a set of selected cultivation conditions(hydroponics,aeroponics,or soil)and abiotic stress treatments(abscisic acid,oxygen deficiency)because a differing stress response could potentially help in explaining the observed higher stress *** apoplastic barriers of poplar roots cultivated in different environments were analyzed by means of histochemistry and gas chromatography and compared to the available literature on monocotyledonous crop ***,dicotyledonous poplar roots showed only a remarkably low induction or enhancement of apoplastic barriers in response to the different cultivation conditions and abiotic stress *** genetic optimization(e.g.,overexpression of biosynthesis key genes)of the apoplastic barrier development in poplar roots might result in more stress-tolerant cultivars in the future.

关键词： Hydroponics Aeroponics Soil Abscisic acid(ABA) Oxygen deficiency Poplar root suberin

Transcriptomic and epigenomic remodeling occurs during vascular cambium periodicity in Populus tomentosa

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Horticulture Research 2021年第1期8卷 1271-1286页

作者： Bo Chen Huimin Xu Yayu Guo paul Grunhofer Lukas schreiber Jinxing Lin Ruili Li Beijing Advanced Innovation Center for Tree Breeding by Molecular Design Beijing Forestry UniversityBeijing 100083China College of Biological Sciences and Biotechnology Beijing Forestry UniversityBeijing 100083China Institute of Tree Development and Genome Editing Beijing Forestry UniversityBeijing 100083China College of Biological Sciences China Agricultural UniversityBeijing 100193China Institute of Cellular and Molecular Botany University of BonnKirschallee 1D-53115 BonnGermany

Trees in temperate regions exhibit evident seasonal patterns,which play vital roles in their growth and *** activity of cambial stem cells is the basis for regulating the quantity and quality of wood,which has received considerable ***,the underlying mechanisms of these processes have not been fully *** we performed a comprehensive analysis of morphological observations,transcriptome profiles,the DNA methylome,and miRNAs of the cambium in Populus tomentosa during the transition from dormancy to *** analysis showed that the active cambial zone exhibited a significant increase in the width and number of cell layers compared with those of the dormant and reactivating ***,we found that differentially expressed genes associated with vascular development were mainly involved in plant hormone signal transduction,cell division and expansion,and cell wall *** addition,we identified 235 known miRNAs and 125 novel *** expressed miRNAs and target genes showed stronger negative correlations than other miRNA/target ***,global methylation and transcription analysis revealed that CG gene body methylation was positively correlated with gene expression,whereas CHG exhibited the opposite trend in the downstream *** importantly,we observed that the number of CHH differentially methylated region(DMR)changes was the greatest during cambium ***,the genes with hypomethylated CHH DMRs in the promoter were involved in plant hormone signal transduction,phenylpropanoid biosynthesis,and plant–pathogen interactions during vascular cambium *** findings improve our systems-level understanding of the epigenomic diversity that exists in the annual growth cycle of trees.

关键词： periodicity finding during

Endoscopic and non-endoscopic approaches for the management of radiation-induced rectal bleeding

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World Journal of Gastroenterology 2016年第31期22卷 6972-6986页

作者： Joseph paul Weiner Andrew Thomas Wong David Schwartz Manuel Martinez Ayse Aytaman David schreiber Department of Radiation Oncology VA NY Harbor Healthcare System Brooklyn Campus Department of Radiation Oncology SUNY Downstate Medical Center Department of Gastro-enterology VA NY Harbor Healthcare System Brooklyn Campus Department of Gastroenterology SUNY Downstate Medical Center

Pelvic radiation is a commonly utilized treatment for malignancy of the genitourinary and lower gastrointestinal tract. Radiation proctitis and the resultant clinical picture varies from asymptomatic to potentially life threatening. Similarly, treatment options also vary greatly, from medical therapy to surgical intervention. Commonly utilized medical therapy includes sucralfate enemas, antibiotics, 5-aminosalicylic acid derivatives, probiotics, antioxidants, short-chain fatty acids, formalin instillation and fractionated hyperbaric oxygen. More invasive treatments include endoscopic-based, focally ablative interventions such as dilation, heater and bipolar cautery, neodymium/yttrium aluminum garnet argon laser, radiofrequency ablation or argon plasma coagulation. Despite its relatively common frequency, there is a dearth of existing literature reporting headto-head comparisons of the various treatment options via a randomized controlled approach. The purpose of our review was to present the reader a consolidation of the existing evidence-based literature with the goal of highlighting the comparative effectiveness and risks of the various treatment approaches. Finally, we outline a pragmatic approach to the treatment of radiation proctitis. In light of the lack of randomized data, our goal is to pursue as least invasive an approach as possible, with escalation of care tailored to the severity of the patient's symptoms. For those cases that are clinicallyasymptomatic or only mildly symptomatic, observation or medical management can be considered. Once a patient fails such management or symptoms become more severe, invasive procedures such as endoscopically based focal ablation or surgical intervention can be considered. Although not all recommendations are supported by level I evidence, reported case series and single-institutional studies in the literature suggest that successful treatment with cessation of symptoms can be obtained in the majority of cases.

关键词： Prostate cancer Radiation therapy Radiation proctitis Radiation proctopathy Medical treatment Endoscopic treatment Hyperbaric oxygen Neodymium/ yttrium aluminum garnet argon laser Argon plasma coagulation

Equal Sex Ratios of a Marine Green Alga, Bryopsis plumosa

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Journal of Integrative Plant Biology 2008年第6期50卷 648-652页

作者： Tatsuya Togashi paul Alan Cox Marine Biosystems Research Center Chiba University Institute for Ethnomedicine Box 3464 Jackson Hole Wyoming 83001 USA Institute for Ethnomedicine Box 3464 Jackson Hole

By finding some important culture conditions as below, we succeeded in experimentally controlling the whole life history of a dioecious marine green alga, Bryopsis plumosa （Hudson） C. Agardh. In this study, we focused on the primary and secondary sex ratios （i.e. at inception and maturity） using these culture techniques. Gametogenesis was induced by culturing haploid gametophytes with Provasoli＇s enriched seawater （PES） medium under a 14：10 h light： dark cycle at 14 ℃. Formed zygotes grew into diploid sporophytes, which were cultured for 3 months with PES medium under a 14：10 h light ： nbsp;dark cycle at 18℃. Then they were transferred into schreiber medium and cultured under a 10：14 h light： dark cycle at 22℃. Within 1 week, zoosporogenesis was observed. Zoospores were released within a couple of days. Each zoospore soon germinated and grew into a unisexual gametophyte. The primary sex ratio was examined in gametophytes that originated from a single sporophyte. The secondary sex ratio was studied in the field. Both were estimated as 1：1. Synchronized meiotic cell divisions might occur during zoosporogenesis dividing each sex-determining factor evenly among zoospores. Given the equal sex ratio at maturity, there seems to be no environmental factor that differentially affects the survival of male or female gametophytes in nature.

关键词： algal culture Bryopsis green algae life history sex ratio.