Week 5

SYCPA Regents Prep Living Environment

Labs 1-5

Week 5 - Labs

This packet provides a review of concepts that may be tested in the NYS Living Environment Regents and is based on the NYS Core Curriculum. This course will cover individual topics over 6 weeks as listed below:
Week Week Week Week Week Week 1 2 3 4 5 6 - Scientific Method - Ecology - Human Body Systems Molecular Biology & Genetics - Labs - Evolution & Human Impact

The order of these topics is chosen based on their average weight in past Living Environment Regents. The individual packets will consist of a review for the specific topic followed by past regents questions.

Good Luck! SYCPA Say Yes Collegiate Prep Academy

Week 5 - Labs

Labs Review
Instrumentation Review Stereo Microscopy Stereo Microscope The image at the left is that of a dissecting or stereo microscope. Notice that it has only two sets of lenses, including the eyepiece. The specimen to be observed is an opaque object (light does not pass through it). The observer sees the surface of the dissection specimen or other specimen being studied. This specimen is placed in a container on the stage of the microscope.

Light Microscopy Biologists use the light microscope to observe microscopic specimens. This microscope is also called the compound microscope is given its name because it has more than two sets of lenses. It has an eyepiece lens (or ocular) and two or more sets of objective lenses (this microscope has three lenses) on a nosepiece that usually revolves. This kind of microscope is also called a light microscope as it requires a source of light to pass through the specimen. The specimen observed with this kind of microscope is usually microscopic and has to be translucent (allows light to pass through it). The specimen to be observed is placed on the stage of this microscope. Parts of the Light Microscope 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. eyepiece or ocular body tube fine adjustment knob nosepiece high power objective low power objective diaphragm mirror (many microscopes have a light instead) base coarse adjustment arm stage clip inclination joint

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Functions of the Light Microscope Parts eyepiece (ocular) - where you look through to see the image of your specimen. body tube-the long tube that holds the eyepiece and connects it to the objectives (not labeled) fine adjustment knob-small, round knob on the side of the microscope used to fine tune the focus of your specimen after using the coarse adjustment knob nosepiece-the rotating part of the microscope at the bottom of the body tube; it holds the objectives high power objective -- used for high power magnification of the specimen (the longer objective lens) low power objective -- used for low power magnification of the specimen diaphragm-controls the amount of light going through to the specimen light or mirror-source of light usually found near the base of the microscope; makes the specimen easier to see base-supports the microscope coarse adjustment knob -- used for focusing on low power arm-part of the microscope that is grasped when one carries the microscope stage clips-shiny, clips on top of the stage which hold the slide in place (The specimen is placed on the stage for viewing.) inclination joint -is used to tilt the microscope

Some Microscope Usage Rules Always carry the microscope with two hands - one on the arm and one underneath the base of the microscope. Hold it up so that it does not hit other objects. Do not touch the lenses. If they are dirty, ask the teacher for special lens paper or ask your teacher to clean the lenses for you. If using a microscope with a mirror, do not use direct sunlight as the light source. Blindness can result. If using a microscope with a light, turn off light when not in use. Notify teacher if a slide or cover slip breaks. Students should not handle broken glass. Always clean slides and microscope when finished. Store microscope set on the lowest power objective with the nosepiece turned down to its lowest position (using the coarse adjustment knob). Cover microscope with dust cover and return it to storage as directed by your teacher.

Other Points About the Compound Microscope 1. Always begin focusing on the lowest possible power. Remember to center the specimen you are observing in the field of view before switching to a higher power. Make certain that you move the objectives away from the specimen when focusing so their is no collision between the objective being used and the slide/cover slip which may damage the objective lens. 2. As you switch from low to high power, the field of view becomes darker. To deal with this the diaphragm needs to be opened to allow in more light. (Frequently on low power the diaphragm needs to be partially closed as it is too bright.)

Week 5 - Labs 3. As you switch from low to high power the field of view becomes smaller. Images viewed under the light microscope are reversed (backward) and inverted(upside down). This is a compound light microscope view of the letter F placed on a slide in its normal position.

Chromatography Paper chromatography is a procedure used to separate substances in a mixture. In the Living Environment/Biology lab, this mixture is usually a solution of liquid plant pigments containing different kinds of chlorophylls and other colored photosynthetic pigments. A small concentrated sample of a mixture is placed on the chromatography paper above the line of a solvent mixture. The paper is contact with a solvent solution at its bottom. This solvent moves through the paper due to capillary action and dissolves the mixture spot. Some parts of the solvent mixture to be separated have a greater attraction for the chromatography paper, so they move a lesser distance, while other parts of the solvent mixture have a lesser attraction, so they move a greater distance up the paper. Paper Chromatography Apparatus

Week 5 - Labs Completed Paper Chromatography of a Plant Pigment

The specific mixture placed on chromatography paper will separate into consistent patterns as long as the same solvent, paper, and amount of time allowed for the separation are not changed.. Different solvents will change the separation pattern of the mixture. Mixtures that are colored can be separated into component colors by paper chromatography. The Rf value of a pigment is a statistic often computed from a chromatography separation. Each component of a solution. Each pigment in the solution will have a specific Rf for the same solvent when the chromatography occurs for a specific length of time.

Calculation of Rf Rf =
Electrophoresis Gel electrophoresis is a procedure used to separate charged molecules of different sizes by passing them through a gel in an electrical field. The gel serves to act as a support for the separation of the molecules of different sizes. The gel is usually composed of a jelly-like material called agarose which is made from seaweed. Electrophoresis Setup

distance the pigment travels from the original spot of solvent distance to the wetting front of the solvent

Week 5 - Labs Molecules such as DNA fragments of different lengths and proteins of different sizes are often separated in the gel. Holes are created in the gel which serve to hold the particular DNA mixtures to be separated. The DNA fragments are then loaded into the wells in the gel.

Separation of DNA

The gel contains very small holes which act to regulate the speed which molecules can move through it based on the size of the molecules. The smaller molecules will move much more easily through the small holes in the gel. As a result, large fragments of DNA lag behind small fragments, thus allowing the experimenter to separate these molecules based on their size.

Sometimes molecular weight markers are electrophoresed along with the specimen, so the experimenter may know the size of the DNA fragment which has been separated. Different individuals or organisms form different banding patterns in the plate when their DNA has been separated. DNA is cut into pieces for separation for electrophoresis by restriction enzymes. These enzymes were originally discovered in bacteria and were used by the bacteria to defend themselves from invasion by other bacteria and viruses.

Some Uses for the Gel Electrophoresis DNA Separation

1. It may be used to determine an individual's genetic relationship to his or her ancestors, as the more closely matched the banding pattern between two individuals, the more closely they will be genetically 7

Week 5 - Labs related. In theory, no two individuals will form the same DNA banding pattern when the electrophoresis is completed. 2. It may be used to identify an individual that have committed crimes based on the ability to match the suspects DNA to evidence which has been collected at a crime scene.

3. It may be used to determine evolutionary relationships

between organisms, as organisms with a closer genetic relationship will form more similar banding patterns. Electrophoresis Setup with Power Supply

Measurement Review Volume Measurement A commonly used instrument to measure liquid volume is the graduated cylinder. This instrument usually measures liquid volume in milliliters (ml). Using a Graduated Cylinder

Week 5 - Labs It is important to remember to read to the bottom of the curved line ormeniscus when measuring solutions involving water or most liquids. The graduated cylinder at the left is divided into increments of 2 ml, so the volume in it is 12 ml. The graduated cylinder on the right is divided into increments of 1 ml, so the volume in it is 16 ml. Mass Measurement The triple beam balance is commonly used to measure mass in the biology lab. This device is named for its three long beams on which sliding bars called riders (or tares) are used to determine the mass of an object placed on its platform. It is very important that the riders on the rear beams are in the notch for the whole number of grams and not in between notches. The front beam is a sliding scale graduated in grams. The rider on this beam can be positioned anywhere on the scale. Masses on a triple-beam balance can be read to tenths of a gram and estimated to hundredths of a gram.

Using the Triple Beam Balance The picture at the upper left shows two different models of triple beam balances commonly used in the biology laboratory. The picture at the lower left shows the measurement of a mass in progress. Without estimation, the mass of the object appears to be 373.3 grams (g).

Length Measurement Most measurements in biology will involve metric units of measurement. It is good to start at a whole number increment that isn't 0. Many times the end of a ruler will be worn away by student/teacher use or is inaccurate due to the manufacturing process. It is important to remember to take away the whole number increment one has moved in on the ruler (in the example below 1 cm) from the measurement obtained. Using a Ruler to Measure Length

Week 5 - Labs Problem: How long is leaf A?

The tip of the leaf is at about 6.5 cm, but note the measurement started at 1 cm. Therefore, Leaf A is 5.5 cm or 55 mm in length.

Microscopic Measurement The magnifying power of most objectives and oculars is engraved on them. On the ocular, the marking can be found on the top edge or on the smooth cylinder that fits inside the body tube; on the objectives, magnification is on the side of the cylinder. For example, a marking "10x" means that the particular lens forms an image ten times larger than the object being viewed. The total magnification of a microscope is equal to the power of the eyepiece (ocular) X power of the objective used. For example, if a student is using a microscope with a 10 X ocular and a 43 X high power objective, the total magnification of the specimen the student is viewing is equal to 10 X 43 or 430 X (times). Formula for Total Microscope Magnification Total magnification = Power of the x eyepiece Power of the objective

The size of a microscopic field of view can be determined on low power using a device called an optical micrometer. An economy version of this can be made by placing a clear metric ruler on the stage of a microscope and using it to estimate the field of view. The light microscope is used to look at cells or other similarly sized microscopic objects, so small units of measure such as millimeters or micrometers are used. It is important to remember that there are 1,000 micrometers in 1 mm (millimeter) and 1000 millimeters in a meter. Finding the Size of a Microscope Field of View In the pictured field of view at the left, it can be observed that there are approximately 3 1/2 divisions equal to a length of 3.5 mm. Therefore this field of view is equal to 3.5 mm or 3,500 micrometers.

Finding the Size of Multiple Cells in a Field of View

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Week 5 - Labs The two cells in this field take up a field of view of one millimeter. Therefore, the size of the specimen is equal to 1 mm/2 cells or 0.5 mm per cell. There is 500 micrometers in 0.5 mm., so the average size of each cell is 500 micrometers.

Estimating Cell Size When the Field of View is Known

It is often difficult to approximate the approximate size of the field of view, but this ameba considered lengthwise appears to occupy approximately 1/3 of the field of view. The field of view in the left image is 3 mm. Given that the ameba in the image takes up about 1/3 of that field, we can find its approximate length by multiplying the 3 mm X 1/3 = 1 mm length or 1,000 micrometers for the approximate length of this ameba. The student is viewing the same ameba in the field of view at the right on a higher power. The field of view gets smaller which makes the ameba appear larger in this field. Indicators and Stains Indicators An indicator is any substance used to assist in the classification of another substance. There are many different kinds of indicators. Some common kinds of indicators used in Living Environment/Biology will be indicated below.

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Week 5 - Labs The pH Scale Acids and bases (alkalis) are common substances studied in science. The pH scale is used to indicate the relative strength of an acid of base. The pH scale goes from 0 to 14. A pH of 7.0 is considered to be neutral. The greater the pH is than 7.0, the more basic the substance is. The lower the pH is below 7.0, the more acidic a substance is. Stomach acid has a pH of approximately 2.0.

Some Common Indicators 1. Litmus paper turns red or a shade of red in acids. Litmus turns blue or a shade of blue in bases. It is important to place a few drops or a small amount of the substance to be tested on the litmus paper when testing it. Do not dip the litmus paper in the substance to be tested. A paper which provides a more specific indication of the pH level of a substance is pH paper. This paper turns different shades of various colors which may be compared to a scale to determine the pH value. 2. Bromthymol blue is an indicator used to show the presence of either carbon dioxide in solution or an acidic solution. Low levels of carbon dioxide or acid will result in the bromthymol blue solution remaining blue, while higher levels of carbon dioxide or acid will result in the bromthymol solution taking on a yellow tint. Frequently this indicator is used in biology labs to indicate photosynthetic activity (solution turns blue as CO2 is used) or respiratory activity (solution turns yellow as CO2 is added to the solution).

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Week 5 - Labs 3. Lugol's solution (which is actually IKI) is a brown solution which turns black in the presence of starches. The test tube at the right shows Lugol's (iodine) solution mixed with a starch suspension.

4. Benedict's solution is used to detect the presence of simple sugars such as glucose. When a simple sugar is mixed in Benedict's solution and heated for a short period of time in a test tube, it goes through a variety of color changes, eventually ending as an orange-red or brick red color. The use of Benedict's solution before and after using it to detect the presence of the simple sugar glucose is shown in the pictures on the right.

Stains Very frequently it is helpful to dye certain cell structures so that they can be seen more clearly. Chemicals that dye parts of cells for this purpose are called stains. Two commonly used stains in the biology laboratory are Lugol's iodine solution and methylene blue. Lugol's solution is a good stain to make the nuclei of plant cells stand out more prominently. It has the unfortunate drawback of killing the cells it used on however. Methylene blue is often used to stain animal cells, such as human cheek cells, to make their nuclei more observable. It is vital dye which does not immediately kill the specimen.

Using Stains in Biology These are plant cells stained withLugol's solution so their nuclei are visible. These are human cheek cells stained with methylene blue solution making their nuclei and outlines much more visible.

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Correctly Staining Specimens

1. A specimen is obtained and placed on the slide with forceps. A cover slip is then lowered on to the specimen from an approximately 45 degree angle gently. This reduces the number of air bubbles the specimen will have. The student then places a drop or two of water on the specimen. 2. The student places a drop of stain beside and under one corner of the cover slip. 3. The student places a towel on the opposite side of the cover slip in the water beside the cover slip. This will draw the stain through the entire specimen in a few seconds without removing the cover slip. This technique will also remove any air bubbles which have formed. The stained specimen may now be observed. Note that this technique can be used to draw salt water or distilled water into a specimen having a cover slip over it without removing the cover slip as well.

Dichotomous Keys A dichotomous key is a sequence of steps that allows the identification of a living thing. The key will consist of a series of choices that lead the user to the correct name of a given item. The term dichotomous means that there will always be two choices in each step of the key until the organism is correctly identified.

Some Key Ideas in Dichotomous Key Construction 1. Use constant characteristics rather than ones that disappear or vary with the season or other environmental factor. 2. Use characteristics which can be directly observed. 14

Week 5 - Labs 3. Use quantitative measurements with an amount or dimension rather than vague terms like "big" and "small." 4. Precede the descriptive terms with the name of the anatomical part to which it applies. Rules to Follow When Using a Dichotomous Key 1. Always read both choices, even if the first seems to be the logical. 2. Understand the meaning of the terms involved in the key. 3. When measurements are given, use a scale to measure the specimen. Do not guess at a measurement. 4. Living things are always variable, so do not base your organism identification in the field on a single observation. Using a Dichotomous Key to Identify an Organism

The example below will illustrate the use of a dichotomous key to identify the unknown creature above. Steps in the Dichotomous Key Taxonomic Key to Stream Water Animals 1. A. With a shell go to 2 B. Without a shell go to 3 2. A. Shell made of two parts held together by a hinge Clam B. Shell made of only one part Snail A. Body flat, oval and brown Water Penny B. Body not exactly like a water penny go to 4 go to ** The creature has 6 jointed legs, so go to # 5. ** This creature does not have an oval body it is long, so go to # 4. Identification Process

** The creature clearly does not have a shell, so go to # 3.

3.

4. A. With six jointed legs 5 B. With more than six jointed legs

go to 12

C. With less than six jointed legs; body often worm-like go to 14 5. A. With two or three thin, ** The creature has three thin tails, so go to # 6. 15

Week 5 - Labs hair-like tails B. Without thin, hair-like tails go to 6

go to 7

6. A. With one hook at the end of each leg; usually with three tails, sometimes only two Mayfly B. With two hooks at the end of each leg; two tails Stonefly 7 A. Body with many long, pointed parts go to 8 B. Body not exactly like this go to 9 8. A. Body brown or black, often very large Hellgrammite B. Body white, yellow or tan; not so large Beetle larva 9. A. Body with hook-like claws at tail end; animal sometimes protected with bits of sand, pebbles or twigs Caddisfly B. Body without hook-like claws go to 10 10 A. Body small, dark, hard and beetle-like Riffle beetle B. Body not exactly like this go to 11 11 A. With 3 wide tails Damselfly B. Without tails, but with three short points Dragonfly 12 A. With two large claws and eight legs; large Crayfish B. Without large claws; smaller go to 13 13. A. Body flattened side to side; usually white Scud B. Body flattened top to bottom; usually gray Sowbug 14. A. Body with very small legs; usually with a head go to 15 B. Body without any legs or head go to 16 15 A. Tail-end of body wider than the

** This organism clearly has three tails and only a single hook at the end of each leg which makes it a Mayfly larva.

We didn't need to go beyond step 6 with the organism we classified above, but some organisms might require the use of many more steps before its proper identification

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Week 5 - Labs other Black fly larva B. Tail-end of body not wider Midge 16. A Body brown, plump, and caterpillar-like Crane fly larva B. Body not exactly like this go to 17 17. A. Body with suckers at each end Leech B. Body without suckers; small, thin and worm-like Aquatic worm

Dissection Anatomical Direction Before beginning a dissection, it is important to have an understanding of some of the basic directional terminology associated with the dissection of specimens. Some of these terms include proximal, which means toward the body, and distal, which means to move away from the body. Other important anatomical directions are indicated below. Key Anatomical Directions

Dissection Safety Proper safety procedures when working with dissection tools and specimens is of greatest importance. Some safety rules to engage in when dissecting specimens are as follows.

Dissection Safety Rules Follow all instructions given by your teacher. Inform your teacher of any illness as a result of exposure to chemicals used in specimen preparation. 17

Week 5 - Labs Avoid contact with preservative chemicals. Rinse the specimens completely before dissection. Know where the eye-wash fountain is if needed. Wear safety goggles to prevent the splashing of any chemicals into the eyes. Properly mount dissection specimens to dissecting pan. Do not dissect a specimen while holding it. Handle scalpel or razor blade (safety edged) with extreme care. Always cut away from your body and away from others. Never ingest specimen parts. Never remove specimens or specimen parts from the classroom -- until the dissection is completed all parts of the dissection must remain within the dissecting pan. Properly dispose of dissected materials. Store specimens in as directed by your teacher. Clean up the work area and return all equipment to the proper place when the dissection is completed. Wash hands after each dissection. Dissection Equipment Dissection Equipment The pictured dissection equipment from left to right is (1.) a teasing or dissection needle which used to pull apart muscle tissue, (2)dissecting scissors which are used to cut through tissue, and (3) a scalpel, which is a knife used to slice through and cut tissue. Plant Dissection Many kinds of flowering plants, such as lilies, daffodils, or tulips are commonly subjects for dissection in biology. The flower is the plant structure specialized for reproduction in advanced plants. The processes of meiosis and fertilization occur in the flower. Some Key Flower structures

petals: colored parts inside the sepals which attract insects sepals: structures which are usually green outside the petals which help to protect the flower 18

Week 5 - Labs stamen: forms the male reproductive organ and consists of an anther and a filament anther: pollen box in which pollen grains are formed containing the genetic material which produces sperm filament: supports the anther pistil or carpel: female reproductive organ which consists of three parts stigma: found at the top of the pistil, is often sticky and hairy adapting it to catch and hold pollen style: tube-like connection between the stigma and the ovary ovary: enlarged part of the pistil attached to the receptacle (stem tip on which the flower rests) and contains the ovules ovules: small white structures within the walls of the ovary which produces the plant egg cells Animal Dissection The dissection of animals is important for many reasons. It helps in the learning about the internal structures of animals. It also allows students to learn how organs and tissues are interrelated. Another purpose of dissection is to allow the comparison of organisms in terms of their organs and relative complexities. While many good simulations of dissections may be observed, it seldom can replace the benefits of the actual participation in an actual dissection. Some common vertebrate organisms dissected in the living environment lab include the frog and the fetal pig. Usually the dissection procedure involves tying the organism down firmly on the dissection pan, cutting the organism open on its ventral side (as pictured below), and pinning its tissues and muscles back to observe its internal organs. Different teachers may have their own preferences in terms of their emphasis on the tissues and organs to be observed in a dissection. Key Internal Organs of the Frog Organ brain heart stomach small intestine liver gall bladder lungs kidneys ureter Body System nervous circulatory digestive digestive digestive (and other systems) digestive respiratory excretory excretory Major Function thinking and coordination of body activities pumps blood through the body stores and begins the chemical digestion of food finishes chemical digestion and absorbs digested nutrients into the blood makes bile, detoxifies poisons, many other functions stores bile from liver for release into small intestine to aid in fat digestion exchanges gases with the external environment (aided by the skin in the frog) filter wastes from the blood carries wastes to the urinary bladder 19

Week 5 - Labs urinary bladder pancreas ovaries testes excretory stores urine before its release from the body produces hormones like insulin which regulate blood sugar, produces pancreatic juice which aids in digestion in the small intestine makes eggs in female frog makes eggs in male frog

endocrine/digestive reproductive reproductive

Frog Internal Anatomy

Provided by Regentsprep.org and The NYS Core Curriculum

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