2D Gel Electrophoresis| Principle & Limitations | 2 Dimensional Gel Electrophoresis | 2DGE | PENS#89
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- เผยแพร่เมื่อ 2 ส.ค. 2024
- 2D Gel Electrophoresis| Principle & Limitations | 2 Dimensional Gel Electrophoresis | 2DGE | PENS#89
#2dgel #2DGE #proteomics
The primary technique for proteomics research is two-dimensional gel electrophoresis, or 2D-PAGE. It uses two different features of proteins to distinguish the complicated mixture of samples. Proteins are distinguished in the first dimension by their pI value, and in the second dimension by their relative molecular weight. Although it was first reported by O'Farrell in 1975, the introduction of immobilised pH gradient strips improved its application and adoption since they provided consistent findings and were easy to handle. Proteins were first visualised using 32P or 35S labelling. SYBRO Ruby, a more delicate approach, has now taken its place. Advances were made in various stages of the 2D-PAGE technology, which can separate up to 10,000 proteins in a single run.
The Lectures are given Dr. Nagendra Singh aiming to spread basic understanding and awareness of biological molecules and reactions. Dr. Nagendra Singh finished his Doctoral and Post Doctoral studies from "All India Institute of Medical Sciences" (AIIMS) New Delhi. Dr. Singh has a vast experience of research and Teaching in the fields of Structure & Molecular Biology and Biochemistry, which also include Biomolecules, Proteomics, Protein Engineering and Drug Design.
Dr. Nagendra Singh currently works at the School of Biotechnology, Gautam Buddha University, Greater Noida, INDIA (www.gbu.ac.in/FacultyProfiles/.... He does research in Molecular Biology, Structural Biology and Biotechnology. His current research interest include 'Structural and functional characterization of proteins involved in RNA metabolism from human and microbial origin. Dr Singh has published over 70 research articles in intentional Journals of good repute. He has over 20 awards and honors on his name. Few of the awards include "National Academy of Sciences of India (NASI, Allahabad) young scientist award", UGC-NET, CSIR-JRF, GATE, Best Research Awards etc. Dr. Singh has participated and presented research work at over 50 national and International conferences world wide.
Thank you sir. You made it crystal clear.💫
I am so glad u like it...
@19.IBT.039 Saiyada Iqra Kamil
Thank you so much for this insightful and detailed video on 2DGE. May I ask you two questions? 1. What is the chemistry behind the formation of the pH gradient in the IPG strips? 2. As SDS completely disrupt the polymeric configuration of a protein, doesn't it affect further downstream analysis like MS methods? It will be helpful if you please answer this.
@Aditi Sarkar ... I will be glad to answer the queries.
1. Buffers are nothing more than a mixture of acid-basic groups. on IPG strips, amphoteric groups are covalently immobilized on a matrix such as polyacylamide to create their concentration gradient.
2. SDS disrupt all structure of proteins except the primary structure (sequence). so after SDS PAGE only sequence of a protein remains, that is what required to identify and further characterization by MS. MS identifies protein by either sequencing in MS/MS or by peptide mass finger printing (PMF). MS does not provide any details about function of 3d structure. so it doesnt matter to MS if protein has lost tertiary or quaternary structure.
I hope it will help u in further understanding. Feel free if any doubt persists.
@@proteinengineering007 Okay! Thank You, Sir. Take homage🙏