title: Polyclonal Antibody Development Against Purified CC-NBS-LRR like Protein Fragment from Mature Lageneria siceraria Seeds and Immunolocalization.
abstract: CC-NBS-LRR (CNL) plant proteins are related with highly conserved family of disease resistance protein distinguished by a coiled-coil domain, which plays an important role in innate immunity. The present study reports the purification and identification of CNL like protein fragment (CNL-LPF) by two step chromatography and matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF/MS), respectively. Furthermore, current study also illustrated the development of polyclonal antibody against purified CNL-LPF, which was used for immunolocalization of CNL-LPF in cytoplasm of cotyledon, using Fluorescence microscopy and Transmission electron microscopy. Lastly, present study also demonstrates in vitro oligomerization of purified CNL-LPF with multiple bands on 4-10 % gradient native-PAGE; each band representing a small fraction of each oligomer population as evident by immunoblots. In conclusion, the current study deals with the purification and polyclonal antibody development against CNL-LPF.
authors.affiliation: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, 110029, India.
authors.forename: Neha
authors.initials: N
authors.lastname: Kumari
authors.affiliation: Translational Health Science and Technology Institute, NCR Biotech Science Cluster, Faridabad, 121001, India.|Department of Biochemistry, All India Institute of Medical Sciences, New Delhi, 110029, India.
authors.forename: Rajesh
authors.initials: R
authors.lastname: Kumar
authors.affiliation: Department of Environmental Sciences, University of Delhi, Delhi, 110007, India.
authors.forename: Vandana
authors.initials: V
authors.lastname: Mishra
authors.affiliation: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, 110029, India. savita11@gmail.com.
authors.forename: Savita
authors.initials: S
authors.lastname: Yadav
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D019662:Cucurbitaceae / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D060467:Disease Resistance / Q000235:genetics; D005260:Female; D015870:Gene Expression; D007106:Immune Sera / Q000737:chemistry; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification*; D007150:Immunohistochemistry; D000070576:NLR Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D057865:Plant Immunity / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D055503:Protein Multimerization; D011817:Rabbits; D012639:Seeds / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D000906:Antibodies; D007106:Immune Sera; D007074:Immunoglobulin G; D000070576:NLR Proteins; D010940:Plant Proteins
keywords: CC-NBS-LRR protein; CNL like protein fragment; Enzyme-linked immunosorbent assay; Immunoblots; Immunolocalization; Lageneria siceraria; Polyclonal antibody