title: An improved immunofluorescence staining method for chloroplast proteins.
abstract: An improved immunofluorescence staining method significantly facilitates the visualization of the subcellular localization of interested proteins in chloroplasts. As an important technical approach, immunofluorescence staining is widely used in the subcellular localization study of interested proteins. During the study of the functions of chloroplast division proteins, immunofluorescence staining was frequently adopted. Previously, a method has been developed to study the localization of a chloroplast division protein, FtsZ. However, it is laborious and time-consuming. In this study, we report a modified immunofluorescence staining method, in which protoplasts were isolated from leaf tissues, and then fixed for immunofluorescence staining. The time of the experiment was significantly reduced to several hours. Furthermore, we used correction pen in the fixation procedure and a new way to coat the slide, which greatly saved the cost of the experiment. With the chloroplast division protein ARC6 as an example, we can get a good fluorescence signal. Moreover, the localization of ARC6 in two chloroplast division mutants, arc3 and arc5, and three other plant species, such as cabbage, radish and pea, was also successfully analyzed with our new method. Overall, the immunofluorescence staining method we reported here is very practical, and it significantly facilitates the visualization of the subcellular localization of interested proteins in plant cells.
authors.affiliation: College of Biological Sciences and Biotechnology, Beijing Forestry University, Beijing, 100083, China.
authors.forename: Yiqiong
authors.initials: Y
authors.lastname: Li
authors.affiliation: College of Biological Sciences and Biotechnology, Beijing Forestry University, Beijing, 100083, China.
authors.forename: Qingqing
authors.initials: Q
authors.lastname: Sun
authors.affiliation: Beijing Key Lab of Bioprocess, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, China.
authors.forename: Yue
authors.initials: Y
authors.lastname: Feng
authors.affiliation: College of Biological Sciences and Biotechnology, Beijing Forestry University, Beijing, 100083, China. liuxiaomin@bjfu.edu.cn.
authors.forename: Xiaomin
authors.identifier: http://orcid.org/0000-0002-9314-8615
authors.initials: X
authors.lastname: Liu
authors.affiliation: College of Biological Sciences and Biotechnology, Beijing Forestry University, Beijing, 100083, China. gaohongbo@bjfu.edu.cn.
authors.forename: Hongbo
authors.initials: H
authors.lastname: Gao
mesh_terms: D000595:Amino Acid Sequence; D000906:Antibodies / Q000378:metabolism; D000941:Antigens / Q000276:immunology; D017360:Arabidopsis / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000378:metabolism*; D060365:Chloroplast Proteins / Q000737:chemistry / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D015151:Immunoblotting; D016415:Sequence Alignment; D013045:Species Specificity; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D000906:Antibodies; D000941:Antigens; D029681:Arabidopsis Proteins; D060365:Chloroplast Proteins
keywords: ARC6; Chloroplast; Immunofluorescence staining; Localization