genes in bottle

8/11/2019 genes in bottle

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UNIVERSITI TEKNOLOGI MARA

FAKULTI KEJURUTERAAN KIMIA

GENETICS LABORATORY (CBE 561)

NAME : NUR NASUHA BINTI HUSA

GROUP : 4A

EXPERIMENT : GENES IN BOTTLE

DATE : 13 MARCH 2014PROG/CODE : EH 222

SUBMIT TO : MOHAMAD SUFIAN BIN SOAIB

No Title Allocated Marks (%) Marks

1 Abstract 5

2 Introduction 5

3 Objectives 5

4 Theory 5

5 Procedures/Methodology 10

6 Apparatus 5

7 Results 10

8 Calculation 109 Discussion 20

10 Conclusion 10

11 Recommendations 5

12 References 5

13 Appendices 5

TOTAL 100

Remarks:

Checked by: Rechecked by:

Date: Date:


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ABSTRACT

This experiment is mainly about the process of extracting the DNA and makes it visible so that

the DNA can be seen by naked eyes. This is important part in a genetic engineering since it is

step of many biotechnological procedures. Basically, this experiment can be done by using any

cells or tissues either fruits or even human beings. This is because, the basic units of life is madeup of cell. It is the simplest form which later can be developed into tissue and organs. Inside the

cell, there will be a nucleus which is the main part that will control the whole cell and it is also

the part that contains DNA. To make the DNA visible, the very first step is the extraction of the

cells then followed by the precipitation.

By this experiment, the DNA of an apple is going to be extracted. As mentioned above, the first

step is the extraction which by means, the DNA of an apple was extracted by using a few

materials which can be found easily. The examples of materials that have been used are liquid

detergent and salt. Both of these compounds play their own roles in DNA extraction.

By the end of the experiment, we can obtain the result by observing whether the DNA thread can

be seen or not. This is the important part as we can conclude whether this experiment succeeded

or not. As for this experiment, it was succeeded but the DNA of the apple cannot be seen clearly

due to some errors. This will later be discussed in the discussion part.

INTRODUCTION

Deoxyribonucleic acid or also well known as DNA is component which can be found in all

living things. This function of DNA is to carry genetic information from parents to theiroffspring. The main fact about the DNA that should be known before proceeding with this

experiment is, it cannot be taken out easily from the nucleus of the cell. This is because, the

nucleus is surrounded by many proteins and also cellular membrane. That is why, the extraction

is important for this experiment. By extraction, all the proteins and the membranes will be

degraded first so that the DNA can be isolated and removed.

As mentioned above, the extraction part involves the usage of some materials which are salt and

also liquid detergent. Before mixing the cell of desired with this solution, the sample must be

crushed mechanically so that the cell wall is slightly break apart the cell wall. Besides, the act of

grinding and smashing the cell will also increase the surface exposure that will also help for thefurther action of extracting the DNA. For this experiment, the apple was crushed by using a

blender as the first step. The mixture containing both salt and liquid detergent then mixed with

the crushed apple.

The cell membrane is made up by two layers of lipids that can be broken by the detergent. This

happen as the detergent comes in contact with the cell, the lipids and protein were captured so


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that the fatty membranes destroyed. By the end of this process, the disruption the cell and

nuclear membrane will occur so that the DNA can be extracted easily. Meanwhile, the function

of salt is to precipitate the carbohydrate and the protein so that these components can be

separated from the desired component which is DNA. It can be said that the mixture of both

detergent and the salt helping this process by poking some holes on the mechanically disrupted

cell.

The next step is the precipitation which involving the addition of cold alcohol into the solution

obtained before. This alcohol will help the segregation and precipitation of the DNA. Basically

the alcohol is used as then DNA soluble in it but the other part of cells are not. The mixture will

be separated into two layers, and the alcohol with mixture of DNA will be on the top of the layer

since it is less dense than water. It is important to use cold alcohol because it will separate the

DNA out of the water solution. Besides, the colder temperature will help in better extraction

since it will slows down the enzyme that breaking the DNA. By the end of this process, the DNA

thread should be possible to see, but a process of centrifugation will produce a better result. This

is because the centrifugation will obtain a very pure sample of DNA.

OBJECTIVES

1. To extract the DNA of the cells.

2. To isolate the DNA of the cells.

THEORY

The DNA structure is long and thick. This DNA contains for different units which is labeled bytheir names. These four units is A for adenine, G for guanine, T for thymine and C for cytosine.

The different arrangement of these four units will then gives codes that will carry information

throughout the system. The arrangements of these all four were also called as genes.

The DNA can be found inside the cell, but still need a few steps to be extracted. By this genes in

bottle experiment, the DNA is extracted and precipitated so that it can be seen by naked eyes.The DNA that is contains in every cells of organism is actually located inside the nucleus.

Within the nucleus, the DNA is copied and coiled tightly together and called as chromosome.

The structure of this DNA is called double helix strand where each of the strand is very thin and

highly coiled together.

As mention above, this experiment is to make the DNA visible. By this laboratory activities that

involves the usage of materials that can be obtain from home, we can easily extract the DNA.

The two main materials that are going to be used is liquid detergent and also table salt whichplay their own different function for this experiment. The solution of detergent is called lysis

buffer which will degrades the layers of phospholipids.


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Meanwhile, the salt that added will cause the DNA to become less soluble in the solution and

eventually will segregate. This is happening due to the positive charge of the salt that causes the

attraction towards the negative charge of DNA backbone due to the its phosphate group.

The last step of this experiment is involving the usage of cold alcohol that called as precipitation.

DNA will be precipitated in this step because of its less solubility compared to the water. Andlastly, the DNA of the cell are able to be seen by naked eyes because it has been released fromcell precipitates due to action of salt and cold alcohol.

APPARATUS AND MATERIALS

Apparatus and materials Function.

1. Glass rod To stir and mix the solution.

2. hot plate To heat the solution.


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3. Beaker. To hold the water and solution.

4. Filter paper To filter the pulp and separate the

solution.

5. Filter funnel To hold the filter paper for

filtration.


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6. Blender To mash the apple into pulp.

7. centrifuge To centrifuge the solution

containing DNA to be more visible.

8. Distilled water. To mix the extraction solution.


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9. Table salt To break the cell wall of the apple.

10.Apple

Fruit used to extract its DNA.

11.Ice cubes Used in precipitation of DNA.

12.Acohol To precipitate the DNA.


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PROCEDURE

Preparation of the extraction solution.

1. 3g of table salt was poured into the 80ml distilled water and mixed together in the 100ml

beaker.2. The solution was stirred until all the salt dissolved.

3. 10ml of the liquid detergent then added into the solution and stirred again.

4. Distilled water then added into the solution to produce 100ml of total volume.

Preparation of the pulp.

1. An apple was clean and rinsed.

2. A knife then used to chop the apple into smaller pieces.

3.

The chopped apple then was transferred into a blender.4. Distilled water was then added together with the apple and blended until pulp was

produce.

DNA extraction and filtration.

1. A glass beaker that filled with tap water was heated for 60 degree Celsius.

2. The pulp of apple was mixed with the extraction solution in another beaker.

3. The solution was stirred using glass rod until it blended together.

4. The beaker containing the solution was put into the glass beaker containing hot water at

60 degree Celsius for 15 minutes.

5.

The beaker then taken out and put in the water with ice cubes for another 5 minutes.

6. After 5 minutes, the beaker was taken out and the solution was prepared for filtration.

7. Filter paper was folded into 4 and the put in the funnel.

8. A small beaker then put under the funnel to collect the filtered solution.

9. The mixed pulp solution then poured into the filter paper.

DNA precipitation and centrifuge.

1. 10ml of the solution collected from the filtration was poured in a test tube.

2. 20ml of icy alcohol then added slowly into the mixture.

3.

The test tube was left to be rest for 5 minutes for precipitation.4. If the DNA cannot be seen clearly, the centrifugation needs to be proceed.

5. The mixed solution and the alcohol was centrifuged at 1000rpm for 5 minutes.


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RESULTS

This is the result that obtained from the experiment and after the centrifugation, but the DNA

does not clearly be seen since there is residue of the apple.


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DISCUSSION

This experiment is expecting to get a DNA from sample of cell that has been use. Supposedly,

the DNA can be seen clearly even without the centrifugation. This experiment has used themixture of salt and also the detergent to remove the cell wall and also the layers of lipid so that

we can extract DNA easily. This is the result of extraction part before the centrifugation step. For

this experiment, we did not know the exact concentration of the liquid detergent that has been

used. This might be one of the reasons that we did not see the DNA clearly. If too concentrated

detergent is use, the DNA might rupture and we could not obtain the expected result. Meanwhile,

too less concentrated detergent will not successfully degrades the lipid layer, so the process of

extracting the DNA affected.

Usually extracting DNA from the fruit cell must be started with crushing the fruit. For example,

the fruit need to be blended first especially for types that is hard and cannot be mashed easily.During crushing of the fruit, usually the peel is not use. This is because, the peels contains lack

of DNA or might contains chemicals that can affect the DNA structure. During this experiment,

the peel of apple was not taken out and all part of it was blend together with high amount of

water. This also might be one of the reasons why the DNA strand cannot be seen clearly. Apart

from that, the DNA of an apple is actually hard to obtain compared to another fruit such as

banana. This is because, the apple that has been use maybe is not in the fresh condition that all

the cell and DNA inside is not abundant as the fresh one.

Filtration is one of the important steps before obtaining the solution containing DNA. This is

because, during the filtration, the residue or waste from the fruit that has been used can beseparated. During the experiment, the suggested filter paper is coffee filter paper because more

DNA can be filtered into the solution. During the experiment, only lab filter paper was used,

which is actually too thick that might cause most of the DNA cannot be filtered. Besides, there is

error occur during the experiment since the residue of an apple is still in the solution and not

fully filtered. After the centrifugation, there is residue of an apple that can be clearly seen. This

residue might cause the DNA to be trapped inside, therefore it cannot be seen clearly.

Centrifugation is the process that needs to do to make the DNA strand more visible if the

precipitation did not gives a clear DNA strand. During the experiment, the centrifugation was use

for 5minutes for 10000rpm. This value might be too harsh and could give some effect towardsthe DNA structure. Supposedly, the centrifugation was started using a smaller rpm value and

repeated with higher value so until the clear visible DNA is obtained.


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CONCLUSION

This experiment is basically divided into few different parts before the DNA can really be

extracted. The experiment started with preparation of extraction solution, precipitation and then

centrifugation. In conclusion, the experiment did succeed although the DNA structure cannot be

seen clearly. The expected outcome is the DNA that is white in color but did not obtainedclearly, this is because the DNA might be hidden inside the residue of the apple. Another several

factors might also cause this to happen and were all discussed in the discussion part.

RECOMMENDATION

1. Before the apple crushed, the peels should be removed first. This is because the peel of an

apple contains thick layer of wax that might affect the DNA of it. Besides, the peels

usually did not have high content of DNA.

2.

During the filtration, the filter paper should not having any holes or wide enough so that

it can filtrate completely. This is important to prevent the residue of the apple are still in

the DNA solution that are going to be centrifuge later.

3. The concentration of the liquid detergent is need to be sure so that the amount of it would

be suitable for the extraction solution. This is important since to high concentration might

rupture the cell together with its DNA.

REFERENCES

Carboni, G. (2007, January). How To Extract DNA From Fuits. Retrieved from Fun Science Gallery:

http://www.funsci.com

Genes in a Bottle Kit DNA Extraction Module. (n.d.). Retrieved from Biotechnology Explorer:

explorer.bio-rad.com

how to extract DNA from a kiwi fruit. (2006, July 1st). Retrieved from the naked scientist:

http://www.thenakedscientists.com/

how to extract DNA from living things. (2014). Retrieved from Genetic Science Learning Centre:

http://learn.genetics.utah.edu/


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APPENDICES

1.The pulp of the apple that have been blended with water.

2. Mixing of extraction solution with the pulp of an apple that has been stirred using glass rod.


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3. Heating of water using hot plate to give before heating the solution.

4. Heating the mixed solution of apple and extracted solution in 60 degree Celsius water for 15

minutes.


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5. The filtration of the solution using filter paper before DNA precipitation method.

6. The apple residue after the filtration process.

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genes in bottle