| Single-step Marker Switching in Schizosaccharomyces pombe Using a Lithium Acetate Transformation Protocol
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Author:
Date:
2016-12-20
[Abstract] The ability to utilize different selectable markers for tagging or mutating multiple genes in Schizosaccharomyces pombe is hampered by the historical use of only two selectable markers, ura4+ and kanMX6; the latter conferring resistance to the antibiotic G418 (geneticin). More markers have been described recently, but introducing these into yeast cells often requires strain construction from scratch. To overcome this problem we and other groups have created transformation cassettes with flanking homologies to ura4+ and kanMX6 which enable an efficient and time-saving way to exchange markers in existing mutated or tagged fission yeast strains.
Here, we present a protocol for single-step marker switching by ...
[摘要] 利用不同的选择标记来标记或突变粟酒裂殖酵母中的多个基因的能力受到仅仅两种可选择标记的历史使用的阻碍，即 +和 kanMX6 ;后者赋予抗生素G418（遗传霉素）抗性。最近已经描述了更多的标记物，但是将它们引入酵母细胞通常需要从头开始施加应变。为了克服这个问题，我们和其他团队已经创建了具有与 + 和 kanMX6 的侧翼同源性的转换盒，这使得能够有效和省时的方式在现有的突变或标记的裂变酵母菌株中交换标记。
＆nbsp;在这里，我们提出了裂殖酵母，粟酒裂殖酵母（Schizosaccharomyces pombe）中醋酸锌转化的单步标记转换方案。以下我们将介绍如何将 ura4 + 标记交换到kanMX6 ， natMX4 或 hphMX4标记，其分别对抗生素G418，海参（clonNAT）或潮霉素B提供抗性。我们还详细介绍了如何交换营养标记的任何 MX 标记，例如 arg3 + ， his3 + ， leu1 + 和 ura4 + 。
背景 这种用于粟酒裂殖酵母的单步骤标记交换方案允许将标记有类型的抗生素标记的任何标记或突变的基因替换为营养标记物（含有arg3 + ， + ，并且已经构建了 ura4 + ）并且为任何 MX交换遗传 ura4 + ...
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| Thioglycollate-elicited Peritoneal Macrophages Preparation and Arginase Activity Measurement in IL-4 Stimulated Macrophages
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Author:
Date:
2015-09-05
[Abstract] Macrophages are an essential cell population of innate immunity that plays important roles in inflammatory processes. Two main different phenotypes have been described with opposing activities: The classically activated macrophages (M1) and the alternatively activated macrophages (M2). Alternative activation of mouse macrophages can be induced by type 2 cytokines such as IL-4 and it is characterized by the regulation of the L-arginine metabolism. M2 macrophages convert arginine to ornithine and urea through the action of Arginase-1. Here we described a method for the isolation of peritoneal macrophages from thioglycollate-elicited mice and alternative activation by stimulation with IL-4. Intraperitoneal injection of thioglycollate elicits large numbers of macrophages into peritoneal ...
[摘要] 巨噬细胞是在炎症过程中起重要作用的先天免疫的必需细胞群体。 已经描述了具有相反活性的两种主要不同表型：经典激活的巨噬细胞（M1）和交替激活的巨噬细胞（M2）。 小鼠巨噬细胞的备选激活可以由2型细胞因子例如IL-4诱导，并且其特征在于L-精氨酸代谢的调节。 M2巨噬细胞通过精氨酸酶-1的作用将精氨酸转化为鸟氨酸和尿素。 在这里我们描述了从巯基乙酸诱发小鼠分离腹膜巨噬细胞和IL-4刺激的替代激活的方法。 腹膜内注射巯基乙酸盐引发大量巨噬细胞进入腹膜腔。
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