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di-Sodium hydrogen phosphate anhydrous

di-Sodium hydrogen phosphate anhydrous

Company: EMD Millipore
Catalog#: 106559
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Protein Localization in the Cyanobacterium Anabaena sp. PCC7120 Using Immunofluorescence Labeling
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Date:
2017-06-05
[Abstract]  Techniques such as immunoflorescence are widely used to determine subcellular distribution of proteins. Here we report on a method to immunolocalize proteins in Anabaena sp. PCC7120 with fluorophore-conjugated antibodies by fluorescence microscopy. This method improves the permeabilization of cyanobacterial cells and minimizes the background fluorescence for non-specific attachments. In this protocol, rabbit antibodies were raised against the synthetic peptide of CyDiv protein (Mandakovic et al., 2016). The secondary antibody conjugated to the fluorophore Alexa488 was used due to its different emission range in comparison to the autofluorescence of the cyanobacterium. [摘要]  诸如免疫荧光的技术被广泛用于确定蛋白质的亚细胞分布。在这里我们报告一种免疫定位蛋白质的方法。 PCC7120通过荧光显微镜检测荧光团结合的抗体。该方法改善了蓝细菌细胞的透化性,并使非特异性附着物的背景荧光最小化。在该方案中,针对CyDiv蛋白质的合成肽(Mandakovic等人,2016)产生兔抗体。使用与荧光团Alexa488缀合的二抗,因为与蓝细菌的自发荧光相比,其发射范围不同。

背景 蓝细菌的免疫荧光已广泛用于细胞鉴定和计数研究(Jin等人,2016)。然而,在蓝细菌中蛋白质的免疫定位尚未有效地实现。定位蛋白质最复发的方法是通过将感兴趣的蛋白质融合到具有不同发射波长的荧光蛋白(绿色荧光蛋白)(与蓝细菌自发荧光相比))以及随后使用落射荧光或共聚焦显微镜(Flores < em,et="" al。,2016;="">
&NBSP;蓝细菌细胞的结构性质是应用免疫荧光技术的主要挑战。它们由内膜(IM),肽聚糖层(PG)和外膜(OM)组成(Rippka,1988; Baulina,2012; Jin等人,2016),附加外多糖层(鞘)。鞘细胞均存在于单细胞和丝状蓝细菌中(Kehr and Dittmann,2015),其厚度,组成和外观取决于生长条件,代谢状态,细胞分化及其他外部和内部参数(Jin et al。 ...

Infection of Nicotiana benthamiana Plants with Potato Virus X (PVX)
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Date:
2016-12-20
[Abstract]  Potato Virus X (PVX) is the type member of Potexvirus genus, a group of plant viruses with a positive-strand RNA genome (~6.4 kb). PVX is able to establish compatible infections in Nicotiana benthamiana, a commonly used host in plant virology, leading to mild symptoms, such as chlorotic mosaic and mottling. PVX has been widely used as a viral vector for more than two decades (Chapman et al., 1992; Baulcombe et al., 1995; Aguilar et al., 2015). It provides a feasible means for the systemic expression in plants of heterologous proteins, such as avirulence factors, proteins with pharmacological properties, etc., (Hammond-Kosack et al., 1995; Gleba et al., 2014), and also as a tool to help decipher the function ... [摘要]  马铃薯病毒X(PVX)是Potexvirus属(一组具有正链RNA基因组(〜6.4kb)的植物病毒)的类型成员。 PVX能够在烟草烟草(Nicotiana benthamiana)中建立相容的感染,这是常用于植物病毒学的宿主,导致轻度症状,如褪色马赛克和斑驳。 PVX已经被广泛用作病毒载体二十多年(Chapman等人,1992; Baulcombe等人,1995; Aguilar等人。它为异源蛋白质植物中的全身表达提供了可行的手段,例如无毒性因子,具有药理学性质的蛋白质,等等(Hammond-Kosack等人, 1995; Gleba等人,2014),并且还作为通过病毒诱导的基因沉默(VACS)(Lacomme和Chapman,2008)帮助解释植物中基因功能的工具。摩擦(A)和农用过滤(B)的两种不同的方案,即允许PVX在N中的有效乘法和传播。本文详细描述本文。摩擦方法需要先前感染的液体,并且通过诱导对叶组织的机械损伤来实现感染,从而允许病毒颗粒穿透植物表面。农杆菌过滤需要先前修饰的农杆菌携带并将PVX序列递送到植物细胞中。将土壤杆菌生长直至饱和,并用注射器将其浸入植物组织中建立感染。这两种方法中的任何一种都可以成功应用,选择应主要基于材料和时间的可用性,但建议在使用嵌合病毒时使用农杆菌滤过。

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