QUESTION

 

Title:

DNA Extraction Practical

Abstract:

Roughly 50 words summarising the aim of the practical and what you achieved (i.e. purified DNA)

Introduction:

A brief section that sets the context of the practical i.e. why is it important to be able to purify DNA? How has the ability to purify DNA contributed to genetic and molecular biology research? Finish with a sentence or two summarising would you did (successfully purified DNA from two different cell types).

Materials and Methods:

This section details what you actually did. It should summarise the process you went through in the practical so that somebody could repeat your work. You may find that the Materials used are most easily presented in a table. Feel free to include photographs of the steps that you took should you want.

Results and Discussion:

This section describes what you actually achieved. You can also include brief discussion on anything that didn’t go as planned, or that you would do differently next time. Please also provide brief answers to the following 4 questions which relate to discussions you had during and after the practical session on day school 4:

1. Why is it important to be able to confirm that the material you isolated is DNA?
2. What other molecules and substances might contaminate your sample?
3. Briefly describe one method that you could use to confirm that the material in your tube is DNA?
4. Provide a brief description of one downstream application or experiment that the DNA could be used for.

References:

List here any references you have used (making sure they are properly cited in the text). Given the nature of this assignment a number of these are likely to be online resources and/or protocol information booklets.

 

 

ANSWER

 

SCIENTIFIC REPORT

Abstract

In this report a description of methods and materials of strawberry and human cheek DNA has been done. Both the samples have been considered in order o highlight the materials and methods for DNA isolation. Along with that importance of identification of DNA is stated along with the downstream applications of isolated DNA. DNA isolation method is described briefly. Importance of the DNA extraction has been also described in this study. From this study, it also can be stated that extraction and evaluation DNA greatly contributes in many fields of science.

Introduction

The ability to extract or purify DNA from a particular source is important as it contributes majorly to understand the genetic cause of a particular disease. Along with that, purification of DNA also helps in the development of diagnostics and drugs. The main purpose of the DNA extraction is for genetic analysis. In addition to that, scientists utilize this extracted DNA for many purposes of their research work such as introduction of DNA to another animal or plant cells. This study focuses on the DNA extraction from two different samples, one is shop bought strawberry, Fragaria x ananassa, and other is the epithelial cells from the inner cheeks of human. After completion of the DNA purification from samples, methods and results could be compared in this study.

Materials and method

Sample one

Materials

• Strawberry

• Zipper lock bag

• Tea strainer

• Chilled ethanol (95%)

• Salt

• Measuring cylinder

• Double distilled water

• Beaker

• Test tubes

• Spatula

• Pineapple juice

• Dish washer liquid

• Pasteur pipette

• Cocktail stick

• Permanent marker

Method

1. A strawberry is cleared and taken in a zip lock bag in properly sealed condition to protect it from contamination.

2. A lump of salt and a pinch of washing liquid are added to it and leave it for one or two minutes in room temperature to let the reaction happen in between the washing liquid and the strawberry juice.

3. After few minutes the entire liquid is filtered and poured in a beaker with the help of tea strainer.

4. A test tube is taken and one fourth of the test tube is filled with pineapple juice and the juice is mixed gently with the strawberry and washing liquid mixture and left for about a minute to react well.

5. The strainer is cleaned and used for the filtering and pouring of the mixture to a test tube and a spatula used to help the mixture to go through the strainer (Peretto et al. 2014).

6. After taking the mixture in the test tube a Pasteur pipette is used to add alcohol in the mixture gently by the side of the test tube until there is a thick layer of one to two centimetres alcohol is visible on the top of the mixture.

7. After one or two minutes a white creamy coagulating substance would be seen in the junction of the mixture and the alcohol layer.

8. Based on research of Christou et al. (2014), a cocktail stick needed to be inserted very gently and by rounding the stick the material would collected and the unravelled white strands of DNA could be seen.

9. Lastly the sample is taken in a container and marked with a permanent marker to be identified easily.

Figure 1: DNA sample of strawberry

Sample two

Materials

• Epithelial cheek cells of human

• Double distilled water

• Test tube

• Lysis buffer

• eppendorf tube

• Protease

• Salt

• Pasteur pipette

• Permanent marker

• Chilled alcohol (95%)

• 50∘ C water bath

• Test tube holder

Method

1. The epithelial layer of the cheek needed to be chewed very carefully without any blood split and be very careful to not eat any food before the work.

2. Need to take some water in the mouth and rinse for at least 30 seconds then take out the water in a test tube and a cap is used to cover the test tube and the test tube would be labelled with a name.

3. Based on techniques explained by Suguna et al. (2014), an eppendorf tube is taken and the lysis buffer is measured 2 ml in the tube and added to the sample containing test tube.

4. After adding the buffer the test tube is shaken in a manner of upside down for at least 5 times.

5. The test tube is now put in a 50∘ C water bath in a test tube rack for 10 minutes to let the protease to react properly.

6. The test tube is taken out and alcohol is added with a help of Pasteur pipette as the sample one and do not let it mix.

7. After 5 minutes of rest the creamy layer of DNA would be visible and shaken gently to help it clump. The sample two DNA is considerably less in amount than the sample one DNA.

Figure 2: Human cheek DNA sample

Description

1. Importance of the ability to identify DNA from sample

DNA extraction involves extraction of nucleic acids from a cell. Along with that, purification of DNA refers to the DNA sample that is free from the protein contamination. It is important to remove as many as substances that could interfere with the downstream processing of the DNA purification. Along with that, identification of DNA sample is important as DNA contributes major role in forensic science, sequencing genomes, detecting bacteria and viruses (whatisbiotechnology.org, 2019). It is important to identify the sample is the DNA in case of the DNA extraction from the strawberry and epithelial cells of the cheeks as both the DNA samples are completely different from each other. In case of the DNA extraction from the Strawberry, it has multiple pairs of chromosome as they are octoploid in nature. On the other hand, epithelial cells of human are diploid.

2. Molecules and substances that might contaminate sample

Mainly a DNA sample can be contaminated with proteins of the source sample or can be contaminated by minerals found in water. In case of strawberry if the sample a chance of enzymes residual can be found as in this case laboratory chemical is not used and only used material is a natural substance. In case of strawberry a chance of pineapple DNA or enzymes can be found in this DNA sample as contaminant. However, in human cheek epithelial sample a chance of contamination by natural minerals like Ca2+, Mn2+ and others is possible (Hussein, et al, 2014). In human mouth there are several enzymes are found that works to make fragments of foods and doing so these enzymes would have these types of minerals that works as coefficients of many enzymes. Mn2+ is a mineral which activates DNA polymerase enzymes that cut DNA in fragments if this mineral resides in the sample then there is no chance of finding any DNA in the sample if the sample contains any of the DNA polymerase enzymes. Other than these microbial contamination can happen during the work of isolation (Glassing, 2016).

3. Briefly describe one method used to confirm that the material in your tube is DNA

There are various processes found by which a DNA can be identified such as RFLP marker technique, PCR analysis and other marker based techniques are used. However, whether a sample is a DNA or not or it is contaminated with protein or other molecules can be identified by the spectrophotometric purity analysis. In this process the sample is inserted in a spectrophotometer and its UV absorbance ratio is calculated in a 260-280 nm wavelength field. If the sample is pure DNA then the ratio would be 1.8 and if it is contaminated with protein or other substances then the ration would be lower than 1.8. According to Kosuri & Church (2014), this experiment is helpful in identifying and checking purity of both strawberry and human cheek DNA. After this test both the DNA can be identified and quantified by the marker techniques (Kosuri & Church, 2014).

4. Provide a brief description of one downstream application or experiment that the DNA could be used for

There are several downstream applications for which DNA used nowadays such as restriction endonuclease digestion, cloning, PCR amplification, sequencing, genotypic and others. In case of human cheek DNA the DNA is 2n but this is a large DNA then it can be cut by restriction enzymes by taking DNA 0.5 to 10 μg and making large or small fragments (Kosuri, & Church, 2014). These fragments can be used in cloning, genotyping and sequencing also. In case of strawberry DNA it also can be digested with the help of restriction enzymes and salts and the fragments would be used in cloning for the help of agriculture industry and also can be used for enhancing strawberry life and other tests (Peretto, et al. 2014).

Conclusion

From the above study, it can be concluded that DNA samples from both samples are different from each other as one is plant cell and the other is animal cell. In both experiments, DNA was extracted using different chemicals but very similar methods. In addition to that, DNA is extracted successfully from the given samples. The strawberry can be reproduced abundantly under domestic condition, but human cell requires an efficient lab-based approach. From the above results and discussion, importance of the ability to identify extracted DNA sample is also can be concluded. In addition to that, other molecules that might contaminate the DNA also have been identified in this study. Brief discussion of results and discussion also helps to understand the difference of the chromosomes within these two samples. It is important to identify the sample is the DNA in case of the DNA extraction from the strawberry and epithelial cells of the cheeks as both the DNA samples are completely different from each other. Along with that, the application of the DNA molecule after extraction has also been analyzed in this study. There are a number of medical applications of this DNA purification like identifying DNA sequences in cancerous tumor that has been analyzed in this study.

Reference List

Christou, A., Georgiadou, E. C., Filippou, P., Manganaris, G. A., & Fotopoulos, V. (2014). Establishment of a rapid, inexpensive protocol for extraction of high quality RNA from small amounts of strawberry plant tissues and other recalcitrant fruit crops. Gene, 537(1), 169-173. Retrieved on 31^st December, 2018. Retrieved from https://www.researchgate.net/profile/Arvind_Singh56/post/Is_there_any_rapid_and_reliable_method_for_high_quality_RNA_extraction/attachment/59d6545d79197b80779abef8/AS%3A521431819329536%401501330441374/download/Gene.pdf

Glassing, A., Dowd, S. E., Galandiuk, S., Davis, B., & Chiodini, R. J. (2016). Inherent bacterial DNA contamination of extraction and sequencing reagents may affect interpretation of microbiota in low bacterial biomass samples. Gut pathogens, 8(1), 24. Retrieved on 31^st December, 2018. Retrieved from https://gutpathogens.biomedcentral.com/articles/10.1186/s13099-016-0103-7

Hussein, A. S., Nalbandyan, A. A., Fedulova, T. P., & Bogacheva, N. N. (2014). Efficient and nontoxic DNA isolation method for PCR analysis. Russian agricultural sciences, 40(3), 177-178. Retrieved on 2^nd January, 2019. Retrieved from https://www.researchgate.net/profile/Ahmad_Hussein8/publication/269371122_Efficient_and_nontoxic_DNA_isolation_method_for_PCR_analysis/links/55f878fc08aec948c47e797f/Efficient-and-nontoxic-DNA-isolation-method-for-PCR-analysis.pdf

Kosuri, S., & Church, G. M. (2014). Large-scale de novo DNA synthesis: technologies and applications. Nature methods, 11(5), 499. Retrieved on 3^rd January, 2019. Retrieved from http://arep.med.harvard.edu/pdf/Kosuri_Church_2014.pdf

Peretto, G., Du, W. X., Avena-Bustillos, R. J., Sarreal, S. B. L., Hua, S. S. T., Sambo, P., & McHugh, T. H. (2014). Increasing strawberry shelf-life with carvacrol and methyl cinnamate antimicrobial vapors released from edible films. Postharvest Biology and Technology, 89, 11-18. Retrieved on 2^nd January, 2019. Retrieved from https://s3.amazonaws.com/academia.edu.documents/44235865/Increasing_strawberry_shelf-life_with_ca20160330-22461-15d91j4.pdf?AWSAccessKeyId=AKIAIWOWYYGZ2Y53UL3A&Expires=1546857047&Signature=SpFrV3zbRGDMJ0rSB402n79Zc%2FI%3D&response-content-disposition=inline%3B%20filename%3DIncreasing_strawberry_shelf-life_with_ca.pdf

poly.edu (2019), DNA Extraction and Gel Analysis, Retrieved on: 3^rd January, 2019, Retrieved From: https://manual.eg.poly.edu/index.php/DNA_Extraction_and_Gel_Analysis

Suguna, S. A. J. J. A., Nandal, D., Kamble, S. U. R. E. S. H., Bharatha, A. M. B. A. D. A. S. U., & Kunkulol, R. A. H. U. L. (2014). Genomic DNA isolation from human whole blood samples by non enzymatic salting out method. Int J pharm pharm sci, 6, 198-199. Retrieved on 30^th December, 2018. Retrieved from https://innovareacademics.in/journal/ijpps/Vol6Issue6/9478.pdf

whatisbiotechnology.org (2019), DNA extraction, Retrieved on: 3^rd January 2019, Retrieved From: http://www.whatisbiotechnology.org/index.php/science/summary/extraction/dna-extraction-isolates-dna-from-biological-material

Looking for best Biology Assignment Help. Whatsapp us at +16469488918 or chat with our chat representative showing on lower right corner or order from here. You can also take help from our Live Assignment helper for any exam or live assignment related assistance.