Genbio.pdf

LABORATORY SAFETY RULES Listen to or read instructuctions carefully. Wear safety goggles to protect your eyes from chemicals, heated materials, or things that might be able to shatter. Notify your teacher if any spills or accidents occur. After handling chemicals, alwa...

LABORATORY SAFETY RULES Listen to or read instructuctions carefully. Wear safety goggles to protect your eyes from chemicals, heated materials, or things that might be able to shatter. Notify your teacher if any spills or accidents occur. After handling chemicals, always wash your hands with soap and water. During lab work, keep your hands away from your face. Always tie your hair. For short hair, use headbands or clips to keep hair away from work setup. Roll up loose sleeves. Know the location of the fire extinguisher, fire blanket, eyewash station and first aid kit. Keep your work area uncluttered. Take to the lab station only what is necessary. Use glasses instead of contact lenses. Never put anything into your mouth during an experiment. No horse playing or playing practical jokes in the laboratory. All things inside the laboratory should be treated as hazardous. Clean up your lab area as courtesy to the following user. GLASSWARE SAFETY  Chipped or cracked glassware should not be used. Broken glassware should not be disposed of in a regular trash can. Dispose of in the proper disposal labeled “sharps/glasses” When pouring liquids, ensure the container you are pouring into rests on a table. If a glassware gets broken, do not try cleaning it yourself. Notify the teacher. LABORATORY HYGIENE Never eat, drink or smoke in a laboratory. Never apply cosmetics Never touch your face, mouth, or eyes Always wash your hands before you leave and especially before eating. When working with equipments — Always do a visual check on electrical equipment before use, looking for obvious wear or defects. PARTS OF A COMPOUND MICROSCOPE Optical components of a compound microscope The term “compound” refers to the microscope having more than one lens. Compound microscopes generate magnified images through an aligned pair of the objective lens and the ocular lens. In contrast, “simple microscopes” have only one convex lens and function more like glass magnifiers. 1. Eyepiece (ocular lens) The eyepiece (or ocular lens) is the lens at the top of a microscope that the viewer looks through. The standard eyepiece has a magnification of 10x. You may exchange with an optional eyepiece ranging from 5x – 30x 2. Binocular body The eyepiece tube It also places the eyepiece and the objective lenses within a distance range, generating in-focus images. For monocular microscopes, there is only one eyepiece tube. Binocular microscopes have two eyepieces that allow you to see with both your eyes. The eyepiece tube is flexible and can be rotated/adjusted to fit the users’ distance between two eyes (interpupillary adjustment). A trinocular microscope has an additional third eyepiece tube for connecting a microscope camera. 3. Objective lenses Objective lenses are the primary optical lenses for specimen visualization on a microscope. Objective lenses collect the light passing through the specimen and focus the light beam to form a magnified image. The objective lenses are the most important parts of a microscope. Most of the time a compound microscope comes with 3 or 4 objective lenses. The most common lenses are: Scanning objective lens (4x) red A scanning objective lens is the lowest magnification of all objective lenses. The name “scanning” objective lens comes from the fact that they provide observers with enough magnification for a wide overview of the slide, essentially a “scan” of the slide. Low power objective lens (10x) yellow The low-power objective lens has more magnification power than the scanning objective lens, and it is one of the most helpful lenses for general viewing purposes. High power objective lens (40x) blue The high-powered objective lens (also known as the “high dry” lens) is ideal for observing fine details within a specimen sample after finding the area of your interest using a low-power objective lens. Oil immersion objective lens (100x) white The oil immersion objective lens provides the most powerful magnification. However, the refractive index of air and your glass slide are slightly different, so a special immersion oil must be added to bridge the gap. Without immersion oil, the 100x lens will not function correctly. The specimen appears blurry, and you will not achieve an ideal magnification or resolution. Objective lenses with higher magnification are usually longer. As a result, the tip of high magnification objective lenses (100x) is very close to the specimen. 4. What does the number on the objective lens mean? The information about an objective lens is labeled on the side. Key information that you should pay attention to is the magnification (i.e., 100x), NA (i.e., 1.25), and required media (i.e., Oil; no label means air). Lenses are color-coded and are interchangeable between microscopes if built to DIN standards. Numerical Aperture (NA) determines the limit of the Resolution that your microscope can achieve. The value of NA ranges from 0.025 for very low magnification objectives (1x to 4x) to as much as 1.6 for high-performance objectives utilizing specialized immersion oils. The higher the NA, the better the Resolution is. 5. Nosepiece Nosepiece is also known as the revolving turret. Nosepiece is a circular structure housing the objective lenses. There are holes where the different objective lenses are screwed in. To change the magnification power, rotate the turret to select different objectives. An audible click identifies the correct position for each lens as it swings into place. When turning the nosepiece, grasped the ring around its edge, not the objectives. Using the objectives as handles can the-center and possibly damage them. Pay special attention to the distance between objectives and slides when you switch from low to high power lenses. 6. Specimen stage The stage is a flat platform that supports the slides. The stage has a hole (called aperture) for the illuminating beam of light to pass through. The stage clips hold the slides in place. If your microscope has a mechanical stage, the slide secured on the slide holder can be moved in two perpendiculars (X – Y) directions by turning two knobs. One knob moves the slide left and right; the other moves it forward and backward. The mechanical stage provides more stable movements of the specimen slide instead of having to move it manually. 7. Coarse and fine focus knobs Two adjustment knobs are used to focus the microscope: a fine focus knob and a coarse focus knob. Both knobs can move the stage up and down. You should use the coarse focus knob to bring the specimen into approximate or near focus. Then you use the fine focus knob to sharpen the focus quality of the image. When viewing with a high-power objective lens, carefully focus by only using the fine knob. These two focus knobs are coaxial, meaning they are built on the same axis with the fine focus knob on the outside. Coaxial focus knobs are more convenient since the viewer does not have to grope for a different knob. 8. Iris Diaphragm Control Arm A lever (or rotating disk) that adjusts the amount of light illuminating the slide. Use just enough light to illuminate the object on the slide and give good contrast. 9. Condenser Condensers are lenses that are used to collect and focus light from the illuminator into the specimen. Condensers can be found under the stage often in conjunction with an iris diaphragm. Condensers are critical to obtaining sharp images at magnifications of 400x and above. 10. Iris Diaphragm Iris Diaphragm is located below the condenser and above the light source. This apparatus can be adjusted to change the intensity and size of the cone of light projected through the slide. Abbe condenser and Iris Diaphragm are essential for high-quality microscopes. Combined, they control both the focus and quantity of light applied to the specimen, respectively 11. Collector lens with field diaphragm The collector/field lenses act to collect light from the light source and focus it at the plane of the condenser diaphragm. The condenser lens acts to project this light, without focusing it, through the sample. 12. Field diaphragm lever controls the amount of light that enters the condenser diaphragm 13. On/Off Switch 14. Base It serves as a support for microscopes. Microscope illuminators are also carried by it. 15. Condenser Control Knob The condenser concentrates and controls the light that passes through the specimen prior to entering the objective. It has two controls, one which moves the Abbe condenser closer to or further from the stage, and another, the iris diaphragm, which controls the diameter of the beam of light. 16. Arm Supports the microscope head and attaches it to the base FORMULAS MICROSCOPY TOTAL MAGNIFICATION eyepiece magnification x objective magnification FIELD OF VIEW Field number/Total Magnification note: millimeter is used for the unit MICROMETRY ESTIMATED SPECIMEN SIZE FOV/No. Of Estimated Specimen Across (Horizontally or Vertically) CALIBRATION FACTOR 1 Stage Micron x 10 um / Ocular Micrometer CALIBRATION FACTOR 2 stage micron / ocular ACTUAL SPECIMEN SIZE Calibrated Factor x Ocular Units Occupied ORGANIC MOLECULES ORGANIC MOLECULES - contain carbon and other elements held together by covalent bonds CARBON - less than 0.03% of earth’s crust - 18% of body weight of humans - most common building block of life  - It is the backbone of amino acids and building blocks of protein - carbon atoms forms two covalent bonds with each oxygen atom MACROMOLECULES - greek word “macros” = long - built (synthesized within the cell itself - smaller macromolecules are called “subunits” - Some are for storing energy within our cells, structural components of cells of extracellular structures such as the bone ORGANIC MOLECULES - broken down by a process called hydrolysis - has four types: carbohydrates, lipids, nucleic acids, proteins DEHYDRATION SYNTHESIS - also called the condensation reaction - each time a subunit is added, the equivalent of a water molecule is removed - synthesizing org molecules needs energy HYDROLYSIS - equivalent of a water molecule is added each time a covalent bond between single subunits in the chain is broken - reverse of dehydration synthesis - ATP is released - use to breakdown molecules of food during digestion - the body obtains much of its energy through hydrolisis such as glycogen CARBOHYDRATES - basic structure of carbohydrates is found in their name - have a backbone of carbon atoms with hydrogen and oxygen attached in the same proportion as they appear in water - carbon is hydrated or combines with water MONOSACCHARIDES - simplest kind of carbohydrate - it means one sugar - consists of carbon, hydrogen, and oxygen in a 1-2-1 ratio  - most common: contains 5 or 6 carbon atoms in either a five or six membered ring - - four of of the MOST IMPORTANT monosaccharides in humans: RIBOSE, DEOXYRIBOSE, GLUCOSE, FRUCTOSE RIBOSE AND DEOXYRIBOSE - Both a five carbon monosacc that are components of nucleotide molecules (the basic building block of the nucleic acids DNA and RNA - difference: deoxyribose has one less oxygen atom than ribose GLUCOSE - a six-carbon monosaccharide - an important source of energy for cells - can be linked together w other molecules by DEHYDRATION SYNTHESIS to form larger carbohydrate molecules OLIGOSACCHARIDES - short strings of monosacc - oligo means a few - linked together by dehydration synthesis SUCROSE - also called a disaccharide ( the sugar formed when two monosaccharides are joined by glycosidic linkage, glycosidic = a type of ether bond that joins a carbohydrate (sugar) molecule to another group, which may or may not be another carbohydrate) - it consists of just two monosacc, glucoe + fructose LACTOSE - glucose + galactose - most common disacc GLYCOPROTEINS - some olugosacc are covalently bonded to certain cell-membrane proteins called glycoproteins - this participate in linking adjacent cells together and in cell-cell recognition and communication POLYSACCHARIDES - stores energy  - during dehydration, thousands of monosacc are joined together into straight or branched chains to form complex carbohydrates called polysacc - poly means many - a convenient way for cells to stockpile extra energy by locking it in bonds of polysaccharide molecule - most important polysacc: consist of long chains of glucose monosacc - animals: glycogen storage polysacc - plants: starch - glucose: any glucose not consumed for energy in short term can be used to create glycogen or lipids and stored within our cells for later use CELLULOSE - a slightly different form of glucose polysacc - plants use it for structural support rather than energy use - nature of chemical bonds in cellulose is such that most animals including humans CANNOT break down cellulose down to glucose units - undigested cellulose in our food contributes to the fiber or roughage - the more rapid excretion of wastes decreases the time of exposure to any carcinogens ( cancer-causing agents) that may be in the waste material LIPIDS - MOST IMPORTANT PHYSICAL CHARACTERISTIC of class of organic molecules - relatively insoluble - most important subclasses of lipids in the body are: trigycerides, phospholipids and steroids TRIGLYCERIDES - called neutral fats or just fats - synthesized from a molecule of glycerol and 3 fatty acids - stored in adipose (fat) tissue - important source of stored energy in the body - most of the energy is located in the bonds between carbon and hydrogen in the fatty acid tails FATTY ACIDS - chains of hydrocarbons, usually 16 to 18 carbons long that end in a group of atoms known as “carboxyl group” - carboxyl = HO —C == O  - fats vary in the length of their fatty acid tails and the ratio of hydrogen atoms to carbon atoms in the tails SATURATED FATS - full complement of two hydrogen atoms for each carbon in their tails - the tails are fairly straight allowing them to pack closely together - generally solid in room temeperature - example: animals fasts uch as butter and bacon grease - a diet in rich saturated fats is thought to contribute o the development of cardiovascular disease UNSATURATED FATS - also called oils - fewer than 2 hydrogen atoms on one or more of the carbon atoms in the tails - as a result, double bonds form beteween adjacent carbons putting KINKS in the tails and preventing the fats from associating closely together - generally liquid in temperature - ex: canola oil PHOSPHOLIPIDS - Modified form of lipid - primary structural component of cell membranes - have a molecule of glycerol as the backbone but they have only two fatty acids tails - The presence of chsrged groups on one end gives the phospholipids a special property: on one end of the molecule is polar and thus soluble in water - whereas the other end is neutral and insoluble in water STEROIDS - composed of four rings - do not look like lipids at all but classified as one because they are relatively insoluble in water - consist of a backbone of three 6-memnered carbon rings and one 5 membered carbon ring to which any number of diff groups may be attached - Exam: cholesterol - cholesterol: essential strcutural component of animal cell membranes and the source of several important hormones including sex hormones estrogen and testosterone PROTEINS  - constructed from long strings of single units called “amino acids” - proteins are constructed from 20 different amino acids - each amino acid has an amino group (a functional group that consists of a single nitrogen atom bonded to two hydrogen atoms) - followed by a carboxyl group on the other, a COH group in the middle and additional designated R ( differentiates amino acids) that represents everything else - some R groups are completely neutral, other are neutral but polar and a few carry a net charge (either positive or negative) - differences in the charge and structure of the amino acids affect the shape and function of the proteins - our bodies can synthesize 11 amino acids if necessary - are formed by DEHYDRATION SYNTHESIS POLYPEPTIDE - polypeptide: a single string of 3 to 100 amino acids - Generally referred to as a protein when it is no longer than 100 amino acids and has a complex structure and a function - some proteins cosist of several polypeptides linked together FUNCTION OF PROTEIN - the function depends on the structure - we can define protein strcuture on at least 3 to four levels PRIMARY STRUCTURE - is represented by its amino acids sequence - in writing, each amino acid is indicated by a three-letter code SECONDARY STRUCTURE - describes how the chain of amino acid is oriented in space - common structure: alpha helix ALPHA HELIX - a right-hand spiral that is stabilized by hydrogen bonds between amino acids at regular intervals BETA SHEET - another common structure stabilized by hydrogen bonds  - flat ribbon - formed when hydrogen bonds join two primary sequences of amino acids side by side TERTIARY - the third level - refers to how the protein twists and folds to form a three-dimensional shape - the protein’s three dimensional shape structure depends in part on its sequence of amino acids, because the locations of the polar and charged groups within the chain determine the locations of hydrogen bonds that hold the whole sequence together QUATERNARY - fourth structure - some proteins refers to how many polypeptide chains make up the protein (if there is more than one) and how they associate with each other NOTE: because the links that determine the secondary and tertiary structures are relatively weak, hydrogen bonds may be broken by nearby charged molecules. This means that the shape of proteins can chance in the presecmce of charged or polar molecules DENATURATION - refers to the permnanent disruption of protein structure leading ti a loss of biological function ENYZMES - regulate the rates of biochemical reactions within cells - Functions a sbiological catalyst CATALYST - a substance that SPEEDS up the rate of chemical reactions without being altered or consumed by the reaction - help biochemical reactions to occur - enzymes serves as a catalyst because as proteins they can change shape - enzymes take one or mroe rwactants and turns them int products HOW IMPORTANT ARE ENYZMES? - the reason we can digest glycogen and starch is that we possess specific enzymes that break the chemical bonds between the glucose monosaccharides in these molecules.  - we cannot digest cellulose because we lack the right enzyme to break it apart. - The changeable shape of an enzyme shows why homeostasis within our cells is so important. - Protein shape is in part determined by the chemical and physical environment inside a cell, including temperature, pH, and the concentrations of certain ions. - Any deviation from homeostasis can affect the shapes and biological activities of dozens of different enzymes and thus alter the course of NUCLEID ACIDS - the nucleic acids, deoxyribonucleic acid (DNA) and ribonucleic acid (RNA). DNA - the genetic material in living things, directs everything the cell does. - It is both the organizational plan and the set of instructions for carrying the plan out. - Because it directs and controls all of life's processes including growth, development, and reproduction, DNA is key to life itself. RNA - a closely related macromolecule, is responsible for carrying out the instructions of DNA and, in some cases, for regulating the activity of DNA itself. - In some viruses, RNA (rather than DNA) serves as the genetic material. NOTE: DNA contains the instructions for producing RNA. RNA contains the instructions for producing proteins. Proteins direct most of life's processes. NUCLEOTIDES - Both DNA and RNA are composed of smaller molecular subunits - consist of - (1) a five-carbon sugar, (2) a single- or double-ringed structure containing nitrogen called a base, and (3) one or more phosphate groups. Eight different nucleotides exist: four in DNA and four in RNA. - Each nucleotide is composed of a five-carbon sugar molecule called deoxyribose (like the five-carbon ribose but missing one oxygen atom), a phosphate group, and one of four different nitrogen-containing base molecules; adenine (A), thymine (T), cytosine (C),  or guanine (G). In a single strand of DNA, these nucleotides are linked together by covalent bonds between the phosphate and sugar groups. - The complete molecule of DNA is actually composed of two intertwined strands of nucleotides that are held togethern by weak hydrogen bonds (Figure 2.24). - The sequence of one strand determines the sequence of the other (they are complementary strands), because adenine bonds only with thymine (two hydrogen bonds are required) and cytosine bonds only with guanine (three hydrogen bonds are required). The code for making a specific protein resides in the specific sequence of base pairs in one of the two strands of the DNA molecule. Notice that the entire genetic code is based entirely on the sequence of only four different molecular units (the four nucleotides). You will learn more about DNA and the genetic code when we discuss cell reproduction and inheritance. A single molecule of DNA carries the code for making a lot of different proteins. It is like an entire bookshelf of information, too big to be read all at once. To carry out their function, portions of the DNA molecule are transcribed into The sugar unit in all four of the nucleotides in RNA is ribose rather than deoxyribose (hence the name ribonucleic acid). One of the four nitrogen-containing base molecules is different (uracil is substituted for thymine). RNA is a single-stranded molecule, representing a complementary copy of a portion of only one strand of DNA. RNA is shorter, representing only the segment of DNA that codes for one or more proteins. NOTE: DNA and RNA are constructed of long strings of nucleotides. Double-stranded DNA represents the genetic code for life, and RNA, which is single-stranded, is responsible for carrying out those instructions. ATP - One additional related nucleotide with an important function is adenosine triphosphate (ATP). ATP is identical to the adenine-containing nucleotide in RNA except that ATP - has two additional phosphate groups - ATP consists of an adenine base, the five-carbon sugar ribose, which together are called adenosine, and three phosphate groups, which are bound to each other and therefore called triphosphate. - the bonds between the phosphate groups contain a great deal of potential energy, ATP is a universal energy source for cells. - Any time a cell needs energy for virtually any function, it can break the bond between the outermost two phosphate groups of an ATP molecule. - The breakdown of ATP produces adenosine diphosphate (ADP) plus an inorganic phosphate group (Pi), plus energy - The energy is available to do work for the cell. Only the bond between the outermost two phosphate groups is broken because it is the weakest bond. - ATP can be replenished by using another source of energy to reattach P; to ADP (Figure 2.26b). The energy to replenish ATP may come from stored energy in the food we eat or from the breakdown of energy storage molecules such as glycogen or fat. You will learn more about ATP as an energy source when we discuss energy utilization by muscles. - 3 Recap ATP is a nearly universal source of quick energy for cells. The energy is stored in the chemical bonds between phosphate groups. =