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Yelena Sterlin Oded Pri‐Tal Gil Zimran Sang‐Youl Park Julius Ben‐Ari Jiorgos Kourelis Inge Verstraeten Maayan Gal Sean R. Cutler Assaf Mosquna 《The Plant journal : for cell and molecular biology》2019,98(5):928-941
Abscisic acid (ABA) receptors belong to the START domain superfamily, which encompasses ligand‐binding proteins present in all kingdoms of life. START domain proteins contain a central binding pocket that, depending on the protein, can couple ligand binding to catalytic, transport or signaling functions. In Arabidopsis, the best characterized START domain proteins are the 14 PYR/PYL/RCAR ABA receptors, while the other members of the superfamily do not have assigned ligands. To address this, we used affinity purification of biotinylated proteins expressed transiently in Nicotiana benthamiana coupled to untargeted LC‐MS to identify candidate binding ligands. We optimized this method using ABA–PYL interactions and show that ABA co‐purifies with wild‐type PYL5 but not a binding site mutant. The Kd of PYL5 for ABA is 1.1 μm , which suggests that the method has sufficient sensitivity for many ligand–protein interactions. Using this method, we surveyed a set of 37 START domain‐related proteins, which resulted in the identification of ligands that co‐purified with MLBP1 (At4G01883) or MLP165 (At1G35260). Metabolite identification and the use of authentic standards revealed that MLBP1 binds to monolinolenin, which we confirmed using recombinant MLBP1. Monolinolenin also co‐purified with MLBP1 purified from transgenic Arabidopsis, demonstrating that the interaction occurs in a native context. Thus, deployment of this relatively simple method allowed us to define a protein–metabolite interaction and better understand protein–ligand interactions in plants. 相似文献
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通过相关数据资料的统计与分析,对2014年江苏省药品注册管理情况进行总结,以提升全省药品注册管理能力,为企业研发、管理人员提供相关参考。 相似文献
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Telomere truncation has been shown to be an efficient technology for the creation of mini-chromosomes that can be used as artificial chromosome platforms for genetic engineering. Artificial chromosome-based genetic engineering is considered to be superior to the existing techniques of randomized gene integration by Agrobacterium or biolistic-mediated genetic transformation. It organizes multiple transgenes as a unique genetic linkage block for subsequent manipulations in breeding. Telomere truncation technology relies on three components: the telomere sequence that mediates chromosomal truncation, a selection marker that allows the selection of transgenic events, and a site-specific recombination system that can be used to accept future genes into the mini-chromosome by gene targeting. These elements are usually pre-assembled before transformation, a process that is both time and labor consuming. We found in this research that the three elements could be mixed to transform plant cells in a biolistic transformation, and produced efficient chromosomal truncations and mini-chromosomes in rice. This system will allow rapid construction of mini-chromosomes with a flexible selection of resistant markers, site-specific recombination systems and other desirable elements. In addition, a rice telotrisomic line was used as the starting material for chromosomal truncations. Mini-chromosomes from the truncations of both the telocentric chromosome and other chromosomes were recovered. The mini-chromosomes remained stable during 2 years of subculture. The construction of mini-chromosomes in rice, an economically important crop, will provide a platform for future artificial chromosome-based genetic engineering of rice for stacking multiple genes. 相似文献
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Constable C 《Bioethics》2012,26(3):157-163
In the United States, the decision of whether to withdraw or continue to provide artificial nutrition and hydration (ANH) for patients in a permanent vegetative state (PVS) is placed largely in the hands of surrogate decision-makers, such as spouses and immediate family members. This practice would seem to be consistent with a strong national emphasis on autonomy and patient-centered healthcare. When there is ambiguity as to the patient's advanced wishes, the presumption has been that decisions should weigh in favor of maintaining life, and therefore, that it is the withdrawal rather than the continuation of ANH that requires particular justification. I will argue that this default position should be reversed. Instead, I will argue that the burden of justification lies with those who would continue artificial nutrition and hydration (ANH), and in the absence of knowledge as to the patient's advanced wishes, it is better to discontinue ANH. In particular, I will argue that among patients in PVS, there is not a compelling interest in being kept alive; that in general, we commit a worse violation of autonomy by continuing ANH when the patient's wishes are unknown; and that more likely than not, the maintenance of ANH as a bridge to a theoretical future time of recovery goes against the best interests of the patient. 相似文献
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