Molecular Biology Department of Medical Laboratory Technology (MLMB-201) Department of Medical Laboratory Technology Faculty of Allied Medical Science Lecturer: Dr. Mohamed Salah El-Din

Intended Learning Outcomes (ILO’s): Molecular biology course provides an overview of the molecular basis to cell structure and function. This course focuses on the structure, biosynthesis and function of DNA and RNA on the molecular level and how these interact among themselves and with proteins. Molecular biology techniques are essential for modern biological and medical research. This course will give you an introduction to DNA and RNA standard techniques. Student will have basic knowledge of: Cell organization. DNA structure and function. DNA Extraction. RNA structure and function. RNA Extraction. Gene expression and protein biosynthesis. Agarose gel electrophoresis for DNA/RNA; and SDS-PAGE for protein. Polymerase Chain Reaction (PCR) – Theory, Types, Application. Gene library and screening DNA sequencing

A Cell organization A1 Prokaryotes A2 Eukaryotes A3 Cellular fractionation

Cells Fundamental working units of every living system. Every organism is composed of one of two radically different types of cells: prokaryotic cells or eukaryotic cells. Prokaryotes and Eukaryotes are descended from the same primitive cell. All extant prokaryotic and eukaryotic cells are the result of a total of 3.5 billion years of evolution.

Cells 70% water 7% small molecules 23% macromolecules salts Lipids Chemical composition-by weight 70% water 7% small molecules salts Lipids amino acids nucleotides 23% macromolecules Proteins Polysaccharides lipids biochemical (metabolic) pathways translation of mRNA into proteins

Life begins with Cell A cell is a smallest structural unit of an organism that is capable of independent functioning All cells have some common features

All Cells have common Cycles Born, eat, replicate, and die

Prokaryotes and Eukaryotes According to the most recent evidence, there are three main branches to the tree of life. Prokaryotes include Archaea (“ancient ones”) and bacteria. Eukaryotes are kingdom Eukarya and includes plants, animals, fungi and certain algae.

Prokaryotes and Eukaryotes, continued Single cell Single or multi cell No nucleus Nucleus No organelles Organelles One piece of circular DNA Chromosomes No mRNA post transcriptional modification Exons/Introns splicing

A1 Prokaryotes Prokaryotes Cell structure Bacterial cell walls

Prokaryotes Prokaryotes are the most abundant organisms on earth. (bacteria and blue-green algae) A prokaryotic cell dose not contain a membrane-bound nucleus. Size from 0.1 to 10 μm

Three basic shapes Spherical(cocci) Rodlike(bacilli) Helically coiled( spirilla) They fall into two groups:the eubacteria and the archaebacteria.

The eubacteria Gram-positive Gram-negative Cyanobacteria (cyano-bacteria) in soil,water and living in or on larger organisms

The archaebacteria Sulfur bacteria Methanogens（methane） They grow in unusual environments: salt brines,hot acid springs,in the ocean depths

Cell structure Each prokaryotic cell is surrounded by a plasma membrane. The cell has no subcellular organelles,only infoldings of the plasma membrane called mesosomes. The deoxyribonucleic aid (DNA) is condensed within the cytosol to form the nucleoid. Some prokaryotes have tall-like flagella.

Cell structure

Bacterial cell walls The peptidoglycan (protein and oligosacca-ride) cell wall protects the prokaryotic cell from mechanical and osmotic pressure. A Gram-positive bacterium has a thick cell wall surrounding the plasma membrane,whereas Gram-negative bacteria have a thinner cell wall and outer membrane,between which is the periplasmic space.

Gram negative cell walls

A2 Eukaryotes Chloroplasts Lysosomes Eukaryotes Plasma membrane Peroxisomes Cytosol Cytoskeleton Plant cell wall Plant cell vacuole Eukaryotes Plasma membrane Nucleus Endoplasmic reticulum Golgi apparatus Mitochondria

Plant cell

Animal cell

Plasma membrane

Cell nucleus

Golgi

Mitochondria structure

Chloroplast

Plant cell wall

vacuole

The cell structure 1

A3 Cellular farctionation Subcellular fractionation Differential velocity centrifugation Equilibrium density-gradient centrifugation Rate-zonal centrifugation Marker enzymes Flow cytometry

Differential velocity centrifugation

Some Terminology Genome: an organism’s genetic material Gene: a discrete units of hereditary information located on the chromosomes and consisting of DNA. Genotype: The genetic makeup of an organism Phenotype: the physical expressed traits of an organism Nucleic acid: Biological molecules(RNA and DNA) that allow organisms to reproduce;

The genome is an organism’s complete set of DNA. More Terminology The genome is an organism’s complete set of DNA. a bacteria contains about 600,000 DNA base pairs human and mouse genomes have some 3 billion. human genome has 24 distinct chromosomes. Each chromosome contains many genes. Gene basic physical and functional units of heredity. specific sequences of DNA bases that encode instructions on how to make proteins. Proteins Make up the cellular structure large, complex molecules made up of smaller subunits called amino acids.

All Life depends on 3 critical molecules DNAs Hold information on how cell works RNAs Act to transfer short pieces of information to different parts of cell Provide templates to synthesize into protein Proteins Form enzymes that send signals to other cells and regulate gene activity Form body’s major components (e.g. hair, skin, etc.)

Assignment: As a part of the semester activity, one student is selected every week to prepare a short seminar about his/her point of interest in one of the lecture topics. That to be discussed and evaluated during the next lecture.