Molecular Cell Biology. Prof. D. Karunagaran. Department of Biotechnology. Indian Institute of Technology Madras
|
|
- Rudolph Waters
- 8 years ago
- Views:
Transcription
1 Molecular Cell Biology Prof. D. Karunagaran Department of Biotechnology Indian Institute of Technology Madras Module 5 Methods in Cell Biology (Methods to Manipulate Protein, DNA and RNA and Methods to Visualize Cells) Lecture 2 Methods to Manipulate Cells and Proteins
2 Isolating Cells Tissues contain a heterogeneous population of cells and these have to be isolated for biochemical studies. Disrupting extracellular matrix, cell-cell interactions and cell-cell adhesions is an important first step to isolate individual cells from tissues. This is done by treatment with proteolytic enzymes such as trypsin and with chelating agents such as ethylene diamine tetra acetic acid. If cells have to be multiplied in large numbers then they can be grown using standard cell culture protocols using appropriate media supplemented with serum and growth factors. Cultures of cells derived directly from tissues obtained from an organism are known as primary cultures. Plant cells can also be grown initially as a mass of undifferentiated cells called callus which can give rise to roots and shoots with appropriate media. Culturing cells Normal animal cells divide only for about times after which they do not divide further and this is called replicative cell senescence. Chromosomes get shortened after each cell division and short DNA sequences called telomeres are necessary to replenish this defect. Since normal cells lack telomerase that takes care of maintaining telomeres the cells are not able to divide forever. If telomerase is made active in these cells they can be grown continuously and are said to be immortalized However, not all human cells can be immortalized by this method of activating telomerase since they may have activated cell cycle checkpoint mechanisms that arrest the cell cycle. Such checkpoint mechanisms can be inactivated by introducing oncogenic viruses or oncogenes and thus the cells can be immortalized. Some rodent cells can inactivate these mechanisms and immortalized cells emerge spontaneously.
3 Immortalized cells prepared from cancer cells are called transformed cells and they can grow for several generations and do not need a solid support to grow. Such cell lines are useful in research and they can be frozen in liquid nitrogen at C and can be thawed again and they still retain their viability. Embryonic stem cells can also proliferate indefinitely and can develop into any other cell type. Specific cell types can be separated from a mixture of cells by fluorescence activated cell sorter. A specific cell type can be labeled with a fluorochrome coupled antibody that binds to its specific surface marker protein. The cells suspended in a fluid are passed through a narrow passage and come in contact with a laser beam and those which emit fluorescent light are now given a charge so that they can be deflected to fall into a collecting tube. Selected cells can be grown in culture or can be used for analysis. Fluorescence activated cell sorter
4 To analyze the biochemical components of cells, they are first subjected to sonication/osmotic shock/grinding. These methods open up the cells breaking them into pieces containing organelles, proteins and other components suspended in a homogenate or extract. The different components of this extract have to be separated by centrifugation. Cellular organelles such as nuclei, mitochondria, and lysosomes differ insize and specific gravity, and these properties are used in centrifugation to separate them. In differential centrifugation different components/organelles of a cell can be pelletted with different speeds of centrifugation. In isopycnic centrifugation, a centrifuge tube is filled with sucrose that is dissolved at different concentrations to produce a density gradient (lower concentration at the top and higher concentration at the bottom. When the cell extract is centrifuged at high speed, individual organelles sediment until their buoyant density exactly matches that in the gradient. Each layer can be collected separately. These methods are highly useful to study the functions of cell organelles/components in isolation. These fractions also serve as starting points for further purification of biomolecules in a cell.
5
6 Purification of Proteins Cell extracts may be processed further with column chromatography to purify the proteins of interest. Column chromatography employs a glass or plastic cylinderical column filled with a permeable solid matrix such as cellulose. Cell extracts in suitable solvents are poured over the column and the various components of a cell are collected as various fractions at the bottom since they travel at different rates through the column.
7 In ion exchange chromatography, positively charged components can be retained in the column if an insoluble matrix that is negatively charged is used and vice versa. Examples include Diethyl aminoethyl cellulose (positively charged) and carboxymethyl cellulose and phosphocellulose (negatively charged) Ionic strength and ph of the solution can be changed systematically to achieve better separation and elution of the required proteins.
8 In gel filtration chromatography cross-linked beads of agarose, dextran or acrylamide are used as matrices. In this method small molecules penetrate into the matrices and stay inside for a long time before they emerge out of the column and hence the large molecules are separated out quickly. Affinity chromatography makes use of the affinity between an enzyme and substrate or a ligand and receptor or an antigen and antibody. The column is filled with matrix that is covalently linked to an enzyme/antibody/ligand. The elution is based on differences in ph or salt concentrations, the conditions in which the affinity can be changed drastically to liberate the binding partners. Silica-based resins are used in high performance liquid chromatography that achieves greater resolution to separate proteins.
9 SDS Polyacrylamide Gel Electrophoresis Proteins have a net positive or negative charge depending on their amino acid composition. Electrophoresis is a popular technique that applies electric field to separate proteins based on their net charge, size and shape. Sodiumdodecyl sulfate (SDS) is a detergent that can give a net negative charge to most of the different types of proteins and thus proteins behave as anions and move towards anode when an electric field is applied. SDS gel electrophoresis uses polyacrylamide gels that are highly cross linked and the consequent pores act as sieves for different proteins and separate them depending on their molecular size apart from the net charge. The disulfide bridges in proteins are broken by the addition of reducing agents such as β-mercaptoethanol and this makes the subunits to separate out and migrate in the gel.
10 The proteins migrate in the gel depending on their size and can be visualized by staining with dyes such as Coomassie Blue. SDS polyacrylamide gel electrophoresis is very useful to separate the subunits of proteins and to assess their molecular weights. Proteins thus separated are usually transferred to a nitrocellulose membrane and then identified using specific antibodies coupled with radioactive/fluorescent/luminescent secondary reagents. This method of transferring the proteins on to a membrane and subsequent detection with labeled antibodies is called Western blotting/immunoblotting. Two-dimensional Gel Electrophoresis Basically the technique separates proteins by electrophoresis in two dimensions. The protein sample is first solubilized with a nonionic detergent together with urea and β-mercaptoethanol to keep their intrinsic charge unchanged. In the first dimension the proteins are separated by isoelectric focusing in a narrow tube of polyacrylamide gel that contains a ph gradient by adding a mixture of special buffers. Proteins have their own isoelectric ph (at this ph there is no net charge) and at this ph in the gradient gel they do not move, thus helping in their separation. The separated proteins are now run in the second dimension using SDSpolyacrylamide gel electrophoresis at right angles to the direction in which isoelectric focusing was carried out. The proteins can then be visualized by various staining techniques In this manner protein having same isoelectric ph can also be separated in the second dimension.
11 Mass spectrometry Proteins from a number of different living organisms have been identified and catalogued and this information can be used to identify an unknown proteins isolated from an organism/cell extract Proteins are first digested to produce smaller peptides and then a laser beam is directed towards the peptides that can be ionized in a closed chamber under vacuum. Under an electric field such ionized peptides will move towards an electrode of opposite charge depending on their net charge and the time taken to reach the electrode gives information about the mass of the peptide. Using a database that contains information about proteins and their peptide fragments under these conditions, the unknown peptide can be precisely identified in most cases. This technique is called matrix-assisted laser desorption time of flight (MALDI- TOF) spectrometry. MALDI-TOF MALDI-TOF can be used to determine the molecular weights of proteins and peptides. As an improvement of MALDI-TOF, collisions with high-energy gas atoms can be used to preferentially cleave the peptide bonds, generating a ladder of fragments, each differing by a single amino acid. A second mass spectrometer can separate these fragments and display their masses. The amino acid sequence of a peptide can then be deduced from these differences in mass and this technique is called MS/MS and can be used to detect post translational modifications of proteins such as phosphorylation/acetylation that change the mass of the protein. Another technique called LC-MS/MS digests the proteins with trypsin first and the short peptides produced are subjected to a series of automated liquid chromatography steps. As a second dimension the separated peptides are analyzed by MS/MS technique.
12 X-Ray Diffraction Three dimensional structures of proteins have been determined mainly by using X-ray diffraction. X-rays are electromagnetic radiations of short wavelength (0.1 nm) A narrow beam of X-rays can be directed on a protein crystal and the scattered rays get reinforced and appear as a diffraction pattern detectable by suitable detectors. The diffraction pattern can give rise to an electron density map that has to be carefully interpreted to get the three dimensional structure of a protein. Proteins are needed in a large quantity and good crystals have to be obtained for this technique to be effective Nuclear Magnetic Resonance (NMR) Not all proteins are crystallized in the laboratory efficiently and hence this technique is quite useful to determine the structures of proteins without crystals as it only requires a concentrated solution of proteins for structural determination. NMR measures the absorbance of radio frequency electromagnetic energy by certain atomic nuclei.
13 This technique depends on the fact that certain atomic nuclei are intrinsically magnetic. Only a limited number of isotopes display this property, called spin. The spinning of a proton generates a magnetic moment. This moment can take either of two orientations, or spin states (called α and β) when an external magnetic field is applied. The energy difference between these states is proportional to the strength of the imposed magnetic field. The α state has a slightly lower energy because it is aligned with this applied field. A spinning proton in an α state can be raised to an excited state (β state) by applying a pulse of electromagnetic radiation (a radio-frequency, or RF, pulse), provided that the frequency corresponds to the energy difference between the α and the β states. In these circumstances, the spin will change from α to β; in other words, resonance will be obtained. Basis of NMR microscopy
14 Study Questions 1. How are proteins purified from cell extracts? 2. How does a typical cell sorter work and what are its applications? 3. Cells can be fixed with a) nitric acid b) Glutaraldehyde c) ethanol d) paraffin 4. Match the following NMR X-ray Diffraction SDS-PAGE Iso electric focusing ph Molecular weight Electron spin Electron scattering 5. For proteins that cannot be crystallized method can be used for obtaining structural information.
Protein immunoblotting
Protein immunoblotting (Western blotting) Dr. Serageldeen A. A. Sultan Lecturer of virology Dept. of Microbiology SVU, Qena, Egypt seaas@lycos.com Western blotting -It is an analytical technique used to
More informationApproaches that can be used to study expression of specific proteins
Approaches that can be used to study expression of specific proteins Receptors and transporters Homogenate binding studies Receptor autoradiography Radiochemical Western blotting Immunohistochemistry/cytochemistry
More informationProteomics in Practice
Reiner Westermeier, Torn Naven Hans-Rudolf Höpker Proteomics in Practice A Guide to Successful Experimental Design 2008 Wiley-VCH Verlag- Weinheim 978-3-527-31941-1 Preface Foreword XI XIII Abbreviations,
More informationMarmara Üniversitesi Fen-Edebiyat Fakültesi Kimya Bölümü / Biyokimya Anabilim Dalı PURIFICATION AND CHARACTERIZATION OF PROTEINS
EXPERIMENT VI PURIFICATION AND CHARACTERIZATION OF PROTEINS I- Protein isolation and dialysis In order to investigate its structure and properties a protein must be obtained in pure form. Since proteins
More informationClassic Immunoprecipitation
292PR 01 G-Biosciences 1-800-628-7730 1-314-991-6034 technical@gbiosciences.com A Geno Technology, Inc. (USA) brand name Classic Immunoprecipitation Utilizes Protein A/G Agarose for Antibody Binding (Cat.
More information(c) How would your answers to problem (a) change if the molecular weight of the protein was 100,000 Dalton?
Problem 1. (12 points total, 4 points each) The molecular weight of an unspecified protein, at physiological conditions, is 70,000 Dalton, as determined by sedimentation equilibrium measurements and by
More information6 Characterization of Casein and Bovine Serum Albumin
6 Characterization of Casein and Bovine Serum Albumin (BSA) Objectives: A) To separate a mixture of casein and bovine serum albumin B) to characterize these proteins based on their solubilities as a function
More informationExpression and Purification of Recombinant Protein in bacteria and Yeast. Presented By: Puspa pandey, Mohit sachdeva & Ming yu
Expression and Purification of Recombinant Protein in bacteria and Yeast Presented By: Puspa pandey, Mohit sachdeva & Ming yu DNA Vectors Molecular carriers which carry fragments of DNA into host cell.
More informationMethods for Protein Analysis
Methods for Protein Analysis 1. Protein Separation Methods The following is a quick review of some common methods used for protein separation: SDS-PAGE (SDS-polyacrylamide gel electrophoresis) separates
More information--not necessarily a protein! (all proteins are polypeptides, but the converse is not true)
00Note Set 5b 1 PEPTIDE BONDS AND POLYPEPTIDES OLIGOPEPTIDE: --chain containing only a few amino acids (see tetrapaptide, Fig 5.9) POLYPEPTIDE CHAINS: --many amino acids joined together --not necessarily
More informationDefinition of the Measurand: CRP
A Reference Measurement System for C-reactive Protein David M. Bunk, Ph.D. Chemical Science and Technology Laboratory National Institute of Standards and Technology Definition of the Measurand: Human C-reactive
More informationProtein purification methods, a practical approach
r i Protein purification methods, a practical approach 2008 AGI-Information Management Consultants May be used for personal purporses only or by libraries associated to dandelon.com network. I Edited by
More informationEZ-Run Protein Gel Solution. EZ-Run Protein Standards. EZ-Run Gel Staining Solution. Traditional SDS-Page Reagents. Protein Electrophoresis
EZ-Run Protein Gel Solution EZ-Run Protein Standards EZ-Run Gel Staining Solution Traditional SDS-Page Reagents Protein Electrophoresis protein electrophoresis Introduction Sodium dodecyl sulfate polyacrylamide
More informationPharmaceutical Biotechnology. Recombinant DNA technology Western blotting and SDS-PAGE
Pharmaceutical Biotechnology Recombinant DNA technology Western blotting and SDS-PAGE Recombinant DNA Technology Protein Synthesis Western Blot Western blots allow investigators to determine the molecular
More informationOptimal Conditions for F(ab ) 2 Antibody Fragment Production from Mouse IgG2a
Optimal Conditions for F(ab ) 2 Antibody Fragment Production from Mouse IgG2a Ryan S. Stowers, 1 Jacqueline A. Callihan, 2 James D. Bryers 2 1 Department of Bioengineering, Clemson University, Clemson,
More informationProtein extraction from Tissues and Cultured Cells using Bioruptor Standard & Plus
Protein extraction from Tissues and Cultured Cells using Bioruptor Standard & Plus Introduction Protein extraction from tissues and cultured cells is the first step for many biochemical and analytical
More informationTECHNICAL BULLETIN. HIS-Select Nickel Affinity Gel. Catalog Number P6611 Storage Temperature 2 8 C
HIS-Select Nickel Affinity Gel Catalog Number P6611 Storage Temperature 2 8 C TECHNICAL BULLETIN Product Description HIS-Select Nickel Affinity Gel is an immobilized metalion affinity chromatography (IMAC)
More informationStructural Bioinformatics (C3210) Experimental Methods for Macromolecular Structure Determination
Structural Bioinformatics (C3210) Experimental Methods for Macromolecular Structure Determination Introduction Knowing the exact 3D-structure of bio-molecules is essential for any attempt to understand
More informationChapter 3 Contd. Western blotting & SDS PAGE
Chapter 3 Contd. Western blotting & SDS PAGE Western Blot Western blots allow investigators to determine the molecular weight of a protein and to measure relative amounts of the protein present in different
More informationElectrophoresis and Electroblotting of Proteins
Electrophoresis and Electroblotting of Proteins The purpose of the next lab exercises will be to study the relative amounts of β-actin in cells of the B-16 melanoma, liver and muscle of mice. Electrophoresis
More informationGuide to Reverse Phase SpinColumns Chromatography for Sample Prep
Guide to Reverse Phase SpinColumns Chromatography for Sample Prep www.harvardapparatus.com Contents Introduction...2-3 Modes of Separation...4-6 Spin Column Efficiency...7-8 Fast Protein Analysis...9 Specifications...10
More informationIntroduction to Proteomics 1.0
Introduction to Proteomics 1.0 CMSP Workshop Tim Griffin Associate Professor, BMBB Faculty Director, CMSP Objectives Why are we here? For participants: Learn basics of MS-based proteomics Learn what s
More informationChemical Basis of Life Module A Anchor 2
Chemical Basis of Life Module A Anchor 2 Key Concepts: - Water is a polar molecule. Therefore, it is able to form multiple hydrogen bonds, which account for many of its special properties. - Water s polarity
More informationThe Techniques of Molecular Biology: Forensic DNA Fingerprinting
Revised Fall 2011 The Techniques of Molecular Biology: Forensic DNA Fingerprinting The techniques of molecular biology are used to manipulate the structure and function of molecules such as DNA and proteins
More informationAiping Lu. Key Laboratory of System Biology Chinese Academic Society APLV@sibs.ac.cn
Aiping Lu Key Laboratory of System Biology Chinese Academic Society APLV@sibs.ac.cn Proteome and Proteomics PROTEin complement expressed by genome Marc Wilkins Electrophoresis. 1995. 16(7):1090-4. proteomics
More informationHiPer Ion Exchange Chromatography Teaching Kit
HiPer Ion Exchange Chromatography Teaching Kit Product Code: HTC001 Number of experiments that can be performed: 5 Duration of Experiment: Protocol: 5-6 hours Storage Instructions: The kit is stable for
More informationWorkshop 14-16 February 2006
Theoretical and practical approaches of Hepatocyte primary culture Workshop 14-16 February 2006 Lecture (2) Disaggregation & purification of target cells Coarse organizer Dr. Abo bakr Mohamed Eltayeb General
More information7 Electrophoresis. µ proportional to Q
7 Electrophoresis Objectives: A) To perform agarose gel electrophoresis of the proteins isolated in last week's experiment and B) to interpret the banding patterns produced by these proteins. Introduction:
More informationBIOLOGICAL MEMBRANES: FUNCTIONS, STRUCTURES & TRANSPORT
BIOLOGICAL MEMBRANES: FUNCTIONS, STRUCTURES & TRANSPORT UNIVERSITY OF PNG SCHOOL OF MEDICINE AND HEALTH SCIENCES DISCIPLINE OF BIOCHEMISTRY AND MOLECULAR BIOLOGY BMLS II / B Pharm II / BDS II VJ Temple
More informationVLLM0421c Medical Microbiology I, practical sessions. Protocol to topic J10
Topic J10+11: Molecular-biological methods + Clinical virology I (hepatitis A, B & C, HIV) To study: PCR, ELISA, your own notes from serology reactions Task J10/1: DNA isolation of the etiological agent
More informationIntroduction to Proteomics
Introduction to Proteomics Why Proteomics? Same Genome Different Proteome Black Swallowtail - larvae and butterfly Biological Complexity Yeast - a simple proteome 6,113 proteins = 344,855 tryptic peptides
More informationserum protein and A/ G ratio
serum protein and A/ G ratio Blood plasma contains at least 125 individual proteins. Serum ( as contrasted with plasma) is deficient in those coagulation protein which are consumed during the process of
More information12/6/12. Dr. Sanjeeva Srivastava. IIT Bombay 2
Dr. Sanjeeva Srivastava IIT Bombay Case studies 2DE: Drug treatment in malaria parasite Plasma proteome analysis of SARS An overview of DIGE technique Case study DIGE: Serum proteome analysis of prostate
More informationBiochemistry Lab SDS PAGE and Western blot General Instructions
Background When an electrical field is applied across a solution, the movement of the charged particles (proteins) is influenced not only by the charge but also the voltage, distance between electrodes,
More informationTwo-Dimensional Gel Electrophoresis (2-DGE)
- Introduction - Sample preparation - First dimension: Isoelectric focusing - Second dimension: SDS-PAGE - Detection of protein spots: staining - Imaging analysis & 2D Gel databases - Spot handling: excision,
More informationWESTERN BLOTTING TIPS AND TROUBLESHOOTING GUIDE TROUBLESHOOTING GUIDE
WESTERN BLOTTING TIPS AND TROUBLESHOOTING GUIDE TIPS FOR SUCCESSFUL WESTERB BLOTS TROUBLESHOOTING GUIDE 1. Suboptimal protein transfer. This is the most common complaint with western blotting and could
More informationDNA SPOOLING 1 ISOLATION OF DNA FROM ONION
DNA SPOOLING 1 ISOLATION OF DNA FROM ONION INTRODUCTION This laboratory protocol will demonstrate several basic steps required for isolation of chromosomal DNA from cells. To extract the chromosomal DNA,
More informationSize Exclusion Chromatography
Size Exclusion Chromatography Size Exclusion Chromatography Instructors Stan Hitomi Coordinator Math & Science San Ramon Valley Unified School District Danville, CA Kirk Brown Lead Instructor, Edward Teller
More informationBiology Behind the Crime Scene Week 4: Lab #4 Genetics Exercise (Meiosis) and RFLP Analysis of DNA
Page 1 of 5 Biology Behind the Crime Scene Week 4: Lab #4 Genetics Exercise (Meiosis) and RFLP Analysis of DNA Genetics Exercise: Understanding how meiosis affects genetic inheritance and DNA patterns
More informationChapter 6. Antigen-Antibody Properties 10/3/2012. Antigen-Antibody Interactions: Principles and Applications. Precipitin reactions
Chapter 6 Antigen-Antibody Interactions: Principles and Applications Antigen-Antibody Properties You must remember antibody affinity (single) VS avidity (multiple) High affinity: bound tightly and longer!
More information2D gel Protocol. 2. Determining Protein Concentration of cell lysates
2D gel Protocol 1. Lysis and Protein Extraction from cells Prepare cell lysates with Trizol extraction by following Kathleen Lyons s protocol: AfCS Procedure Protocol PP00000155, Version 1, 05/12/03 (Ref.1).
More informationDiscontinuous native protein gel electrophoresis
Discontinuous native protein gel electrophoresis Michael Niepmann and Junfeng Zheng Institute of Biochemistry Friedrichstrasse 24 Faculty of Medicine, JustusLIebigUniversity 35392 Giessen, Germany In this
More informationDP419 RNAsimple Total RNA Kit. RNAprep pure Series. DP501 mircute mirna Isolation Kit. DP438 MagGene Viral DNA / RNA Kit. DP405 TRNzol Reagent
Overview of TIANGEN Products DP419 RNAsimple Total RNA Kit DP430 RNAprep pure Kit(For Cell/Bacteria) DP315/DP315-R TIANamp Virus DNA/RNA Kit DP431 RNAprep pure Kit (For Tissue) Silica-membrane Technology
More informationDetection of proteins by lithium dodecyl sulphate polyacrylamide gel electrophoresis
Detection of proteins by lithium dodecyl sulphate polyacrylamide gel electrophoresis During electrophoretic measurements a mixture of compounds in solution is taken into a chamber, two electrodes are joined
More informationATOMS AND BONDS. Bonds
ATOMS AND BONDS Atoms of elements are the simplest units of organization in the natural world. Atoms consist of protons (positive charge), neutrons (neutral charge) and electrons (negative charge). The
More informationPROTEIN SEQUENCING. First Sequence
PROTEIN SEQUENCING First Sequence The first protein sequencing was achieved by Frederic Sanger in 1953. He determined the amino acid sequence of bovine insulin Sanger was awarded the Nobel Prize in 1958
More information13C NMR Spectroscopy
13 C NMR Spectroscopy Introduction Nuclear magnetic resonance spectroscopy (NMR) is the most powerful tool available for structural determination. A nucleus with an odd number of protons, an odd number
More informationChapter 2 Antibodies. Contents. Introduction
Chapter 2 Antibodies Keywords Immunohistochemistry Antibody labeling Fluorescence microscopy Fluorescent immunocytochemistry Fluorescent immunohistochemistry Indirect immunocytochemistry Immunostaining
More informationLab 5: DNA Fingerprinting
Lab 5: DNA Fingerprinting You are about to perform a procedure known as DNA fingerprinting. The data obtained may allow you to determine if the samples of DNA that you will be provided with are from the
More informationTroubleshooting Guide for DNA Electrophoresis
Troubleshooting Guide for Electrophoresis. ELECTROPHORESIS Protocols and Recommendations for Electrophoresis electrophoresis problem 1 Low intensity of all or some bands 2 Smeared bands 3 Atypical banding
More informationNO CALCULATORS OR CELL PHONES ALLOWED
Biol 205 Exam 1 TEST FORM A Spring 2008 NAME Fill out both sides of the Scantron Sheet. On Side 2 be sure to indicate that you have TEST FORM A The answers to Part I should be placed on the SCANTRON SHEET.
More informationDNA Separation Methods. Chapter 12
DNA Separation Methods Chapter 12 DNA molecules After PCR reaction produces many copies of DNA molecules Need a way to separate the DNA molecules from similar sized molecules Only way to genotype samples
More informationPRESTWICK ACADEMY NATIONAL 5 BIOLOGY CELL BIOLOGY SUMMARY
Name PRESTWICK ACADEMY NATIONAL 5 BIOLOGY CELL BIOLOGY SUMMARY Cell Structure Identify animal, plant, fungal and bacterial cell ultrastructure and know the structures functions. Plant cell Animal cell
More informationBiology 309 Lab Notebook
Name: Biology 309 Lab Notebook This is a guided lab notebook for you to keep well-organized notes about procedures and record experimental data for experiments as they are performed. It is guided because,
More informationBNG 331 Cell-Tissue Material Interactions. Biomaterial Surfaces
BNG 331 Cell-Tissue Material Interactions Biomaterial Surfaces Course update Updated syllabus Homework 4 due today LBL 5 Friday Schedule for today: Chapter 8 Biomaterial surface characterization Surface
More informationChapter 3. Protein Structure and Function
Chapter 3 Protein Structure and Function Broad functional classes So Proteins have structure and function... Fine! -Why do we care to know more???? Understanding functional architechture gives us POWER
More informationBrochure More information from http://www.researchandmarkets.com/reports/2172993/
Brochure More information from http://www.researchandmarkets.com/reports/2172993/ Proteomics Today. Protein Assessment and Biomarkers Using Mass Spectrometry, 2D Electrophoresis,and Microarray Technology.
More informationGenomic DNA Extraction Kit INSTRUCTION MANUAL
Genomic DNA Extraction Kit INSTRUCTION MANUAL Table of Contents Introduction 3 Kit Components 3 Storage Conditions 4 Recommended Equipment and Reagents 4 Introduction to the Protocol 4 General Overview
More informationEvery time a cell divides the genome must be duplicated and passed on to the offspring. That is:
DNA Every time a cell divides the genome must be duplicated and passed on to the offspring. That is: Original molecule yields 2 molecules following DNA replication. Our topic in this section is how is
More information2007 7.013 Problem Set 1 KEY
2007 7.013 Problem Set 1 KEY Due before 5 PM on FRIDAY, February 16, 2007. Turn answers in to the box outside of 68-120. PLEASE WRITE YOUR ANSWERS ON THIS PRINTOUT. 1. Where in a eukaryotic cell do you
More informationLecture 10 PROTEIN ISOLATION
Lecture 10 PROTEIN ISOLATION 1 Protein isolation Outlines: assay; homogenization; fractionation; centrifugation; quantitation; chromatography, electrophoresis; sequencing; people. 2 Nobel Prize Winners
More informationPrinciples and Applications of Proteomics
Principles and Applications of Proteomics Why Proteomics? 2-DE Overview Sample preparation 1 st & 2 nd dimension seperation Data Analysis Sample preparation for Mass Spectrometry Mass Spectrometry MALDI-TOF,
More informationCHEMISTRY STANDARDS BASED RUBRIC ATOMIC STRUCTURE AND BONDING
CHEMISTRY STANDARDS BASED RUBRIC ATOMIC STRUCTURE AND BONDING Essential Standard: STUDENTS WILL UNDERSTAND THAT THE PROPERTIES OF MATTER AND THEIR INTERACTIONS ARE A CONSEQUENCE OF THE STRUCTURE OF MATTER,
More informationPlant Genomic DNA Extraction using CTAB
Plant Genomic DNA Extraction using CTAB Introduction The search for a more efficient means of extracting DNA of both higher quality and yield has lead to the development of a variety of protocols, however
More informationRecommended Procedures for the Extraction of RNA. Jan Pedersen USDA, APHIS, VS, National Veterinary Services Laboratories, Ames, IA 50010
Recommended Procedures for the Extraction of RNA Jan Pedersen USDA, APHIS, VS, National Veterinary Services Laboratories, Ames, IA 50010 RNA Extraction Isolates RNA from other cellular components in the
More informationPrentice Hall. Chemistry (Wilbraham) 2008, National Student Edition - South Carolina Teacher s Edition. High School. High School
Prentice Hall Chemistry (Wilbraham) 2008, National Student Edition - South Carolina Teacher s Edition High School C O R R E L A T E D T O High School C-1.1 Apply established rules for significant digits,
More informationMolecular Biology Techniques: A Classroom Laboratory Manual THIRD EDITION
Molecular Biology Techniques: A Classroom Laboratory Manual THIRD EDITION Susan Carson Heather B. Miller D.Scott Witherow ELSEVIER AMSTERDAM BOSTON HEIDELBERG LONDON NEW YORK OXFORD PARIS SAN DIEGO SAN
More informationPractical Cell Analysis
Practical Cell Analysis Dimitri Pappas Dept of Chemistry & Biochemistry, Texas Tech University, USA WILEY A John Wiley and Sons, Ltd, Publication Contents Preface Acknowledgments xiii xix 1 Getting Started
More informationAn In-Gel Digestion Protocol
An In-Gel Digestion Protocol This protocol describes the digestion of a protein present in an SDS-PAGE gel band with trypsin. The band can be taken from either a 1D or 2D electrophoresis gel. Reagents
More informationLAB TOPIC 4: ENZYMES. Enzyme catalyzed reactions can be expressed in the following way:
LAB TOPIC 4: ENZYMES Objectives Define enzyme and describe the activity of enzymes in cells. Discuss the effects of varying enzyme concentrations on the rate of enzyme activity. Discuss the effects of
More informationELUTION OF DNA FROM AGAROSE GELS
ELUTION OF DNA FROM AGAROSE GELS OBTECTIVE: To isolate specific bands or regions of agarose-separated DNA for use in subsequent experiments and/or procedures. INTRODUCTION: It is sometimes necessary to
More informationStudent name ID # 2. (4 pts) What is the terminal electron acceptor in respiration? In photosynthesis? O2, NADP+
1. Membrane transport. A. (4 pts) What ion couples primary and secondary active transport in animal cells? What ion serves the same function in plant cells? Na+, H+ 2. (4 pts) What is the terminal electron
More informationYour partner in immunology
Your partner in immunology Expertise Expertise Reactivity Reactivity Quality Quality Advice Advice Who are we? Specialist of antibody engineering Covalab is a French biotechnology company, specialised
More informationIntroduction to mass spectrometry (MS) based proteomics and metabolomics
Introduction to mass spectrometry (MS) based proteomics and metabolomics Tianwei Yu Department of Biostatistics and Bioinformatics Rollins School of Public Health Emory University September 10, 2015 Background
More informationAutomation in Genomics High-throughput purification of nucleic acids from biological samples. Valentina Gualdi Operational Scientist PGP
Automation in Genomics High-throughput purification of nucleic acids from biological samples Valentina Gualdi Operational Scientist PGP OVERVIEW Nucleic acid purification technologies general aspects Genomic
More informationRubisco; easy Purification and Immunochemical Determination
Rubisco; easy Purification and Immunochemical Determination Ulrich Groß Justus-Liebig-Universität Gießen, Institute of Plant Nutrition, Department of Tissue Culture, Südanlage 6, D-35390 Giessen e-mail:
More information1.1.2. thebiotutor. AS Biology OCR. Unit F211: Cells, Exchange & Transport. Module 1.2 Cell Membranes. Notes & Questions.
thebiotutor AS Biology OCR Unit F211: Cells, Exchange & Transport Module 1.2 Cell Membranes Notes & Questions Andy Todd 1 Outline the roles of membranes within cells and at the surface of cells. The main
More informationIntroduction to flow cytometry
Introduction to flow cytometry Flow cytometry is a popular laser-based technology. Discover more with our introduction to flow cytometry. Flow cytometry is now a widely used method for analyzing the expression
More informationMass Spectrometry Based Proteomics
Mass Spectrometry Based Proteomics Proteomics Shared Research Oregon Health & Science University Portland, Oregon This document is designed to give a brief overview of Mass Spectrometry Based Proteomics
More informationLab 10: Bacterial Transformation, part 2, DNA plasmid preps, Determining DNA Concentration and Purity
Lab 10: Bacterial Transformation, part 2, DNA plasmid preps, Determining DNA Concentration and Purity Today you analyze the results of your bacterial transformation from last week and determine the efficiency
More informationA Novel Bioconjugation Technology
A Novel Bioconjugation Technology for Assay Development and More! Presentation overview Who we are Solutions we provide for our customers Solulink s technology Linking system The Solulink advantage Applications
More informationIntroduction to Flow Cytometry
Outline Introduction to Flow Cytometry Basic Concept of Flow Cytometry Introduction to Instrument Subsystems Daisy Kuo Assistant Product Manager E-mail: daisy_kuo@bd.com BDBiosciences Application Examples
More informationIntroduction to Flow Cytometry
Introduction to Flow Cytometry presented by: Flow Cytometry y Core Facility Biomedical Instrumentation Center Uniformed Services University Topics Covered in this Lecture What is flow cytometry? Flow cytometer
More informationPHYSIOLOGY AND MAINTENANCE Vol. II - On The Determination of Enzyme Structure, Function, and Mechanism - Glumoff T.
ON THE DETERMINATION OF ENZYME STRUCTURE, FUNCTION, AND MECHANISM University of Oulu, Finland Keywords: enzymes, protein structure, X-ray crystallography, bioinformatics Contents 1. Introduction 2. Structure
More informationAdvances in Biopharmaceutical and Vaccine Manufacturing Plants
Hitachi Review Vol. 62 (2013), No. 4 267 Advances in Biopharmaceutical and Vaccine Manufacturing Plants Sei Murakami, Dr. Eng. Haruo Suzuki Keisuke Shibuya, Dr. Sc. OVERVIEW: The development of innovative
More informationAnatomy and Physiology Placement Exam 2 Practice with Answers at End!
Anatomy and Physiology Placement Exam 2 Practice with Answers at End! General Chemical Principles 1. bonds are characterized by the sharing of electrons between the participating atoms. a. hydrogen b.
More informationOrganic Molecules of Life - Exercise 2
Organic Molecules of Life - Exercise 2 Objectives -Know the difference between a reducing sugar and a non-reducing sugar. -Distinguish Monosaccharides from Disaccharides and Polysaccharides -Understand
More informationNuclear Magnetic Resonance notes
Reminder: These notes are meant to supplement, not replace, the laboratory manual. Nuclear Magnetic Resonance notes Nuclear Magnetic Resonance (NMR) is a spectrometric technique which provides information
More informationRESTRICTION ENZYME ANALYSIS OF DNA
University of Massachusetts Medical School Regional Science Resource Center SUPPORTING MATHEMATICS, SCIENCE AND TECHNOLOGY EDUCATION 222 Maple Avenue, Stoddard Building Shrewsbury, MA 01545-2732 508.856.5097
More informationObjectives: Vocabulary:
Introduction to Agarose Gel Electrophoresis: A Precursor to Cornell Institute for Biology Teacher s lab Author: Jennifer Weiser and Laura Austen Date Created: 2010 Subject: Molecular Biology and Genetics
More informationX Ray Flourescence (XRF)
X Ray Flourescence (XRF) Aspiring Geologist XRF Technique XRF is a rapid, relatively non destructive process that produces chemical analysis of rocks, minerals, sediments, fluids, and soils It s purpose
More information1. The diagram below represents a biological process
1. The diagram below represents a biological process 5. The chart below indicates the elements contained in four different molecules and the number of atoms of each element in those molecules. Which set
More informationIntroduction to Proteomics
Introduction to Proteomics Åsa Wheelock, Ph.D. Division of Respiratory Medicine & Karolinska Biomics Center asa.wheelock@ki.se In: Systems Biology and the Omics Cascade, Karolinska Institutet, June 9-13,
More informationHow To Make A Tri Reagent
TRI Reagent For processing tissues, cells cultured in monolayer or cell pellets Catalog Number T9424 Store at room temperature. TECHNICAL BULLETIN Product Description TRI Reagent is a quick and convenient
More informationINSTRUCTIONS 56-1190-98. Edition AC
Sephacryl S-100 High Resolution Sephacryl S-200 High Resolution Sephacryl S-300 High Resolution Sephacryl S-400 High Resolution Sephacryl S-500 High Resolution INSTRUCTIONS Sephacryl High Resolution chromatography
More informationGenomic DNA Clean & Concentrator Catalog Nos. D4010 & D4011
Page 0 INSTRUCTION MANUAL Catalog Nos. D4010 & D4011 Highlights Quick (5 minute) spin column recovery of large-sized DNA (e.g., genomic, mitochondrial, plasmid (BAC/PAC), viral, phage, (wga)dna, etc.)
More informationLa Protéomique : Etat de l art et perspectives
La Protéomique : Etat de l art et perspectives Odile Schiltz Institut de Pharmacologie et de Biologie Structurale CNRS, Université de Toulouse, Odile.Schiltz@ipbs.fr Protéomique et Spectrométrie de Masse
More informationVCE CHEMISTRY 2008 2011: UNIT 3 SAMPLE COURSE OUTLINE
VCE CHEMISTRY 2008 2011: UNIT 3 SAMPLE COURSE OUTLINE This sample course outline represents one possible teaching and learning sequence for Unit 3. 1 2 calculations including amount of solids, liquids
More informationLecture Overview. Hydrogen Bonds. Special Properties of Water Molecules. Universal Solvent. ph Scale Illustrated. special properties of water
Lecture Overview special properties of water > water as a solvent > ph molecules of the cell > properties of carbon > carbohydrates > lipids > proteins > nucleic acids Hydrogen Bonds polarity of water
More informationSubject Area(s) Biology. Associated Unit Engineering Nature: DNA Visualization and Manipulation. Associated Lesson Imaging the DNA Structure
Subject Area(s) Biology Associated Unit Engineering Nature: DNA Visualization and Manipulation Associated Lesson Imaging the DNA Structure Activity Title Inside the DNA Header Image 1 ADA Description:
More information