Study of Mitosis in Onion root tips.
All living organisms grow, differentiate differentiate and reproduce by cell division. There are two types of cell division namely, mitosis mitosis and meiosis. Mitosis occur in somatic cells and is known as duplication division. It results in 2 daughter cells with same number of chromosomes. Each mitotic cell division has two events. . !aryokinesis" #ivision of the nucleus. 2. $ytokinesis" #ivision of the cytoplasm. The events that occur during karyokinesis ka ryokinesis are . %rophase" Appearance of chromatids and disappearance of nuclear membrane. 2. Metaphase" &pindle fibres are prominent, holding the chromatids at the e'uatorial plane. (. Anaphase" Movement of chromatids towards poles by splitting at the centromere. ). Telophase" *ormation of daughter nuclei. The meristematic cells of plants provide suitable material for studying mitosis. The root apices of onion is used in the e+periment.
Aim To prepare a stained s'uash of the the onion root tip. Materials re'uired -nion bulbs, eaker, water, water, tooth pick or pins, specimen tubes, a pair of scissors, forceps, forceps, mounting needle, methyl alcohol, acetic acid, hydrochloric acid/ acetocarmine, distilled water, spirit lamp, slides, coverslip, blotting paper, compound microscope. %rocedure Take medium si0ed onion bulbs and trim off the old roots from its base with a sharp pair of scissors. %lace the bulbs on the beakers b eakers filled with water with the base touching the water. 1eave it undisturbed for the new roots to develop. hen the roots start growing, cut 3 mm of the tips of roots and put them in a fi+ative made up of ( glacial acetic acid and ethanol. This will help preservation of the dividing tissues. The cutting should be done in the morning between 4 and 5 AM. 6emove the root tips from the fi+ative and soften them by heating with 78$l at 9:o$ for 3 minutes. 6emove the tips and wash them in distilled water. %lace a drop of freshly prepared aceto carmine on the slide and place the root tip on it. ;ently warm the stain on a spirit lamp by holding the slide much above the flame. %lace a coverslip on the specimen and with the backside of the needle prepare a s'uash by gently running the needle in one direction only. -bserve the specimen under low and under high power and record your observation. -bservation , 1 or I shaped. Telophase Te lophase The spindle disappears and the chromosomes assemble at the polar region. 7uclear membrane reappears. $ytokinesis The two nucleated cells with cell plate at centers are observed. %recautions . ase of onion bulbs to be in contact with water while growing roots. 2. 6oot tips should be fi+ed in the morning. (. The slide must not touch the flame while warming. ). The rolling of needle should be done gently in one direction.
Study of the action of salivary amylase on starch
En0ymes are biocatalysts re'uired for metabolic activities in living organisms. The digestive en0ymes belong to the group namely, 8ydrolases. &aliva is secreted by the salivary glands. It is viscous, co lourless with a specific gravity of .::( and the p8 ranges from 9.2 to 4.). It contains the en0yme salivary amylase or ptyalin that a cts on starch and converts it into simple disaccharide like maltose. salivary &tarch maltose, isomaltose,limit de+trins. amylase Aim To study the effect of salivary amylase on starch. Materials re'uired oiling tube, test tubes, test tube stand, test tube holder, measuring cylinder, dimpled tile, pipette, funnel, beakers, droppers, cotton wool, burner, thermometer, water bath, iodine solution, soluble starch, sodium chloride, enedict/s reagent, *ehling/s A and reagent, distilled water, warm water, labels and dimple tile. %rocedure 6epeat the same every 2 minutes and observe the colour change of the iodine solution. 7ote the time when the colour change does not occur and record the observations. This is known as the achromic point. -bservation The iodine in the dimple will change to blue"black initially. The reaction may not occur after 9 or 5 minutes in the e+perimental %reparation of ? starch solution Take gm of soluble starch in a beaker and add : ml distilled water. &tir well to make a paste. Add @: ml of hot distilled water and stir it. oil the solution to completely dissolve the starch. $oo l and filter the solution. %reparation of ? sodium chloride solution Take : gm of 7a$1 and make :: ml of the solution with distilled water to get ? 7a$l solution. 7a is the cofactor of en0yme amylase. $ollection of saliva 6inse the mouth clean with water. &ip about 2: ml of luke"warm water a nd keep it in the mouth for 3 to : seconds. ;ently collect the same in a boiling tube. This gives the correct dilution of the saliva. 1abel the tube. %reparation of substrate tubes Take 2 clean test tubes and label A"E+perimental and "$ontrol. Take ml of starch solution and I ml of 7a$l solution in both the test tubes A and . %lace both in a hot water bath that is maintained at (4o$ for about : to 3 minutes. Add ml of saliva solution to A and ml of distilled water to . Take a dimple tile with 9 dimples. 1abel the dimples in linear order A" : minutes, A"2min , A"( minB.. and " :min, "2 min, "( minBB and so on. Add 2 drops of iodine solution to the labelled dimples. Take 2 drops each from A and and add to the dimples labelled A": min and ":min respectively. 6ecord the colour turning blue black in a tabular column. mi+ture. The control will remain the same. 7ote the time of the achromic point. $onfirm the reaction.by confirmatory tests. $onfirmatory Test To 2 ml of the en0yme"substrate mi+ture A, add 2 ml of enedict/s solution. oil and observe the colour change. 6epeat the test with *ehling/s A and solution. 6epeat the tests with the content in . 6ecord your observation. Inference The mi+ture when tested with the reagents change from green to yellow to orange to brick red confirming the breakdown of starch to simple sugars. The starch is broken down to maltose at the CCCCCCCCth minute. The achromic point is CCCCCCCCCCCCC. DDDDDDDDDDDDDDDDDDDDDDDDDDDDDD #raw a tabular column with three columns, titled" time, e+perimental tube and control on the left page and enter your recordings.
Study the effect of different temperature on the action of salivary amylase on starch
En0ymes typically operate best in a relatively narrow range of environmental conditions. Many of the en0ymes in our bodies work best at body temperature. At significantly lower temperatures the substrate molecules do not have enough kinetic energy for the reaction to take place even in the presence of the en0yme. At body temperatures significantly higher than normal, the en0yme will not work well because •
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8ydrogen bonds are easily disrupted by increasing temperature. This, in turn, may disrupt the shape of the en0yme so that its affinity for its substrate diminishes. The kinetic energy from the molecules in the solution containing the en0yme is so high, that the en0ymes shape is pulled apart to the point that the en0yme is not able to properly function.
8uman en0ymes generally work best at our body temperature"(4o$. Aim To study the effect of different temperature on the action of salivary amylase on starch. Materials re'uired oiling tube, test tubes, test tube stand, test tube holder, measuring cylinder, pipette, funnel, beakers, droppers, cotton wool, burner, thermometer, water bath, iodine solution, soluble starch, sodium chloride, enedict/s reagent, *ehling/s A and reagent, distilled water, warm water, ice cubes, labels and dimple tile. %reparation of substrate tubes Take 9 clean test tubes and label A, A2, A("E+perimental and , 2, ("$ontrol. Take ml of starch solution and I ml of 7a$l solution in all the 9 test tubes. %repare three water bath" 5:o$, (4:$ and one with ice cubes. %lace A and in water bath 5:o$, A2 and 2 in (4:$ and A( and ( on ice cubes for about : to 3 minutes. Add ml of saliva solution to A series and ml of distilled water to series. %rocedure Take a dimple tile with 9 dimples. 1abel them in linear order A, A2, A( and , 2, (. Add 2 drops of iodine solution to the labelled dimples. Take 2 drops each from A and and add to the dimples labelled A and respectively. 7ote the time as : minute. $lean the dimple tiles and repeat the same every ( or ) minutes and observe the colour change of the iodine solution. 7ote the time when the colour change does not occur and record the observations. This is known as the achromic point. -bservation The iodine in the dimple will change to blue"black initially. The reaction may not occur after 9 or 5 minutes in the e+perimental mi+ture A2. The A, A( and the control serieswill remain the same. 7ote the time of the achromic point. $onfirm the reaction. Inference The mi+ture when tested with the reagents change from green to yellow to orange to brick red confirming the breakdown of starch to simple sugars. The breakdown of starch to educing sugar occurred only in A2 as it provides the optimum temperature for the reaction. 8igh temperature"5:o$, will denature the en0yme and low temperature will inhibit the en0yme action. The optimum temperature for salivary amylase is (4:$.
Study the effect of different pH on the action of salivary amylase on starch
The activity of en0ymes is strongly affected by changes in p8. Each en0yme works best at a certain p8, its activity decreasing at values above and below that point. E+amples the protease pepsin works best as a p8 of F2 Gfound in the stomachH while the protease trypsin is inactive at such a low p8 but very active at a p8 of 5 Gfound in the small intestine as the bicarbonate of the pancreatic fluid neutrali0es the arriving stomach contentsH. • •
$hanges in p8 alter the state of ioni0ation of charged amino acids that may play a crucial role in substrate binding andor the catalytic action itself. Aim To study the effect of different p8 on the action of salivary amylase on starch. Materials re'uired oiling tube, test tubes, test tube stand, test tube holder, measuring cylinder, pipette, funnel, beakers, droppers, cotton wool, burner, thermometer, water bath, iodine solution, soluble starch, sodium chloride, enedict/s reagent, *ehling/s A and reagent, distilled water, warm water, p8 tablets, labels and dimple tile. %reparation of substrate tubes Take 9 clean test tubes and label A, A2, A("E+perimental and , 2, ("$ontrol. Take ml of starch solution and I ml of 7a$l solution in all the 9 test tubes. %lace all the test tubes in a hot water bath that is maintained at (4o$ for about : to 3 minutes. Add p82 GacidicH to A and , p84 GneutralH to A2 and 2, p8: GalkalineH to A( and ( Add ml of saliva solution to A series and ml of distilled water to series. %rocedure Take a dimple tile with 9 dimples. 1abel them in linear order A, A2, A( and , 2, (. Add 2 drops of iodine solution to the labelled dimples. Take 2 drops each from A and and add to the dimples labelled A and respectively. 7ote the time as : minute. $lean the dimple tiles and repeat the same every ( or ) minutes and observe the colour change of the iodine solution. 7ote the time when the colour change does not occur and record the observations. This is known as the achromic point. -bservation The iodine in the dimple will change to blue"black initially. The reaction may not occur after 9 or 5 minutes in the e+perimental mi+ture A2. The A, A( and the control series will remain the same. 7ote the time of the achromic point. $onfirm the reaction. Inference The mi+ture when tested with the reagents change from green to yellow to orange to brick red confirming the breakdown of starch to simple sugars. The breakdown of starch to reducing sugar occurred only in A2 as it provides the optimum p8 for the reaction. The optimum p8 for salivary amylase is p8 4 GneutralH.
