DNA Extraction in Homelabs

Theertha · February 13, 2024, 3:21pm

DNA extraction at CUBE HomeLab. Join whenever n discussion in Context2curriculm group and in Chatshaala.Photo shared by @Arunan sir .

A great discussion about the extraction of DNA from Chicken brain to Strawberry .Thanks to @⁨Sakshi⁩ @⁨~Akanksha Joshi⁩ @purva @Shahin @⁨Amirtha CUBE⁩ @⁨Chithra Mam⁩ and others.

What was the logic behind using soap to wash hands during Covid times ?

Can we do complex DNA Extraction Procedure in homelabs?

How soap helping in breaking the cell membrane of Corona virus@⁨~Akanksha Joshi⁩ @⁨Sakshi⁩ @purva and others?

Can we extract DNA from Strawberry ? or any other common things you can think of @⁨Sakshi⁩ @⁨~Akanksha Joshi⁩ @⁨Himanshu Joshi⁩ @⁨Chithra Mam⁩ and others.How about trying an attempt to extract DNA in our Homelabs? @⁨Abhijeet CUBE⁩ @⁨Amirtha CUBE⁩ @⁨Sakshi⁩ and others.

With only a few household items, you can extract DNA from strawberries just about anywhere! Extracting DNA from strawberries is super easy, allowing you to learn about genomics. Join NHGRI Director Eric Green and social media specialist Jenny Montooth as they show you step-by-step how to extract DNA from strawberries using materials available in your kitchen!"

https://www.genome.gov/about-genomics/teaching-tools/strawberry-dna-extraction#:~:text=With%20only%20a,in%20your%20kitchen!

A fruit with seeds outside like strawberry

Date - 06/02/24, Time - 9:21, location - Delhi , Data collecter - Thikan sahu from chaitanya college pamgarh.Photo shared by @Thikan .

"“Simple, good old-fashioned soap and water is exceptionally effective at removing the coronavirus from your hands - more so than harsh chemicals or disinfectants that can also damage the skin,” adds Dr Corbett. "
How soap kills the COVID-19 virus | Coronavirus (COVID-19) | Queen's University Belfast

““In the case of the virus, this coat is actually a bilayer – consisting of two layers of these phospholipids with tails facing in, and heads facing out. When we wash our hands with soap and water, the tails of the soap molecules start to look for an area where there isn’t any water, and as they do so, they start to surround the virus particles.”
How soap kills the COVID-19 virus | Coronavirus (COVID-19) | Queen's University Belfast.

ISOLATION OF DNA from Chicken Brian ?? Why @⁨Sakshi⁩ and others are interested in chicken brain to isolate DNA !? Why not chicken muscle/blood !? @⁨Enas CUBE⁩ @⁨Amirtha CUBE⁩ @⁨Sakshi⁩ @⁨~Akanksha Joshi⁩ &others

Why soap came in picture in COVID times? Why are we using soap to wash out hands? @⁨Enas CUBE⁩ @⁨~Charvi⁩ @⁨Amirtha CUBE⁩ @⁨Chithra Mam⁩ @⁨Sakshi⁩ @⁨~Akanksha Joshi⁩ & others

How to isolate DNA ? How to break cell membrane and nuclear membrane? @⁨Sakshi⁩ @⁨Enas CUBE⁩ @⁨Amirtha CUBE⁩ & others

What is the function of soap in cleaning oils from hand !? @⁨Enas CUBE⁩ @⁨~Charvi⁩ @⁨Amirtha CUBE⁩ & others

https://www.youtube.com/watch?v=d2IBcOI22uI.

Follow up discussion:
[07/02, 12:44 pm] Enas CUBE: Dische’s diphenylamine reagent consists of diphenylamine, glacial acetic acid, sulfuric acid, and ethanol.[1]

When heated with DNA, it turns blue in the presence of DNA. A more intense blue color indicates a greater concentration of DNA.

[07/02, 12:46 pm] Enas CUBE: Can we think of any alternative to confirm the presence of DNA? @⁨Arunan sir ⁩ @⁨Theertha⁩ @⁨Sakshi⁩ @⁨shalini CUBE⁩ @⁨~Charvi⁩ @⁨Chithra Mam⁩ mam @⁨Batul Ma’am⁩ @amritha @⁨~Akanksha Joshi⁩
[07/02, 4:17 pm] Theertha: What may be the function of each of the components in Dische’s reagent ? How may these components detect DNA?

[07/02, 4:18 pm] Theertha: The acid converts deoxyribose to a molecule that binds with diphenylamine to form a blue substance.
All acids can convert deoxyribose to a molecule that binds with diphenylamine?? @⁨Enas CUBE⁩ @⁨Amirtha CUBE⁩ & others

The reagent does not interact with RNA, so can be used to distinguish DNA from RNA.

The muscle cell, also known as the myocyte is the smallest subunit of all muscular tissues and organs throughout the body. It is here in the myocyte, where the physiological steps of muscle contraction and where the pathophysiology of numerous muscular diseases takes place. https://www.ncbi.nlm.nih.gov/books/NBK544225/#:~:text=The%20muscle%20cell%2C%20also%20known,numerous%20muscular%20diseases%20takes%20place.

[07/02, 9:49 pm] Theertha: Multinucleation is one mechanism adopted by cells to generate and sustain large cell sizes. Muscle cells are one of the largest cell types, which are formed by fusion of mononucleated myoblasts and contain up to several tens (invertebrates) to several hundred (vertebrates) nuclei. Myonuclei are typically positioned at the cell’s periphery, and are distributed to maximize internuclear distance.
Mechanical positioning of multiple nuclei in muscle cells - PMC.

[07/02, 9:49 pm] Theertha: At higher salt concentrations, protein solubility usually decreases, leading to precipitation; this effect is termed salting-out ((Green and Hughes, 1955). Salts that reduce the solubility of proteins also tend to enhance the stability of the native conformation. In contrast, salting-in ions are usually denaturants.salt precipitation of proteins - Google Search

Extraction of DNA in Homelabs: can we extract DNA from banana? How are we going to do this @⁨Abhijeet CUBE⁩ @⁨Amirtha CUBE⁩ @⁨Enas CUBE⁩ @⁨Chithra Mam⁩ @⁨Himanshu Joshi⁩ @⁨Sakshi⁩ & others .

@⁨Sakshi⁩ did you extracted DNA from Chicken Brian? @⁨Sakshi⁩ @⁨~Akanksha Joshi⁩ @purva and others ? Why are we not interested in chicken muscle to extract DNA?@⁨Abhijeet CUBE⁩ @⁨Enas CUBE⁩ @⁨Aditya Joshi⁩ @⁨Amirtha CUBE⁩ & others

Following summary written by @Enas

@⁨Sunitha Mam⁩ mam has given introduced the CUBE in a meeting and about various model organism.
From Covid-19 to nowadays journey.

• A cell ( animal ) has 2 barrier: cell membrane and nuclear membrane when we have To isolate DNA out of it we have to break these two barrier.

• How can we break these barriers? Using detergent example SDS ( sodium dodecyl sulphate).

• Meaning of Hydrophilic and hydrophobic.

• What is covalent bond and non-covalent bond?

• What a detergent and cell membrane is made of ? ( hydrophobic(lipid part) and hydrophilic(inorganic or charged molecules)) .

• When we mix oil in water what happens and why so ?
• When glucose mixed with what happes? Does glucose molecules polar?

• What will happen if a phospholipid monolayer is been kept in water?
How phospholipid bilayer of cell membrane is formed?
@⁨Arunan sir ⁩ @⁨Theertha⁩@⁨Sakshi⁩@⁨Sunitha Mam⁩ mam @⁨Chithra Mam⁩ mam @⁨~Dhanraj⁩

https://www.scienceabc.com/humans/why-is-dna-white.html

Amrita’s plan is to extract DNA from Banana.

• What things are common between plant and animal cell?
• What is the colour of DNA? If white what could be the reason ?

We saw a vedio regarding DNA extraction from strawberry.
• Strawberry have 8 copies of each chromosome. So it’s a Octapoloid.
• What is polypoloidy ?
• what could be the reason of polypoloidy?
• How octapoloidy occurs in strawberry?
• What is genome and subgenome ?

Then disscussed about mitosis and meiosis.
• tetrapoloidy can occur when protein which are participating in cell cycle is basically gets mutated which then may cause the diplod gametes to be formed which give rise to tetrapoloidy.

What is apomixis?
⁩ @⁨Chithra Mam⁩ mam @⁨Theertha⁩ @Amritha @⁨Himanshu Joshi⁩ sir

Polypoloidy is more found in plants example strawberry and wheat etc but in animals case it is less common. In case of drosophila the salivary gland is only polypoloid and not who organism is polypoloid , what could be the reason?
@⁨Theertha Cube⁩ @⁨~Batul Pipewala⁩ mam @⁨~Chitra⁩ mam @⁨~Abhijeet Singh⁩ @⁨~Himanshu Joshi⁩ sir

Theertha · February 13, 2024, 3:23pm

Today’s discussion was between Enas(Bsc. student from AND college), *who is going to extract DNA in Homelabs," and Itika (Bsc. Student from IIS Jaipur) who extracted DNA from Home

The interesting thing was that both the *procedures explained by Enas & Itika were the same.

The only difference is Enas planning to take 2 teaspoons of detergent, while Itika took 3 teaspoons.

Does that matter?
Please describe the setup @⁨Enas CUBE⁩ @itika

Procedure:

Chosen fruit: Banana
Smashing using a plastic cover.
Adding 1 teaspoon of salt, 2 teaspoons of detergent, and a small amount of water.
Filter using cotton cloth.
The filtrate is now treated with sanitizer.

Result:
White cloudy DNA can be seen. @⁨Aditya Joshi⁩ @⁨Enas CUBE⁩ @⁨Chithra Mam⁩ @⁨Himanshu Joshi⁩ & others .

Later, we came to the details of each step: @⁨Enas CUBE⁩ @⁨Chithra Mam⁩ @⁨Himanshu Joshi⁩ @⁨Aditya Joshi⁩ @itika & others

Why detergents:
Detergents can break down the cell membrane and nuclear membrane of plant cells.

Why salt:
Salt neutralizes the charges on the phosphate backbone of DNA molecules. Salt is NaCl; when in contact with water, it ionizes to Na+ and Cl-. Water molecules bond with Na+ and Cl-. These Na+ ions can bind to the phosphate backbone of DNA. As a result, the DNA molecules tend to clump together with salt, preventing precipitation. But at the same time, proteins are getting precipitated.
How? Any comments?
What concentration of salt can cause the precipitation of proteins?@⁨Aditya Joshi⁩ & others

Why alcohol:
Alcohol can remove water molecules from the surroundings of DNA and leads to aggregation/precipitation.

How can we explain each ingredient of our DNA Extraction method by asking why and how?
@⁨Enas CUBE⁩ @⁨Aditya Joshi⁩ @⁨shalini CUBE⁩ @⁨Abhijeet CUBE⁩ & others

https://www.youtube.com/watch?v=gnhUh6qVD5Y. Video shared by @⁨Himanshu Joshi⁩ . What we meant by Salt in & Salt out?
Is Salt in/Salt out applicable in DNA extraction methods?@⁨Aditya Joshi⁩ @⁨Enas CUBE⁩ & others

Theertha · February 13, 2024, 3:28pm

DNA EXTRACTION IN HOMELAB by Amritha .

[12/02, 12:11 pm] Amirtha CUBE: DNA EXTRACTION FROM GRAPES.

REQURIMENTS:1.one glass of water.
2. Two steel bowl.
3.salt.
4.dish wash.
5.plastic cover.
6.one spoon.
Amritha.k sn college alathur.
Time:12:00pm.
Place:Palakkad.
Date:12.2.24.

Theertha · February 13, 2024, 3:33pm

Video from Amritha,about the Extraction of DNA in Homelab.

Theertha · February 13, 2024, 3:45pm

Congratulations @⁨Amirtha CUBE⁩ for made an attempt to extract DNA from your homelabs.

Do you happy about the result?

Do you think we can improve? @⁨Amirtha CUBE⁩ ?

Why are we adding detergents into the DNA extraction method?

How detergents works @⁨Amirtha CUBE⁩ @⁨Aditya Joshi⁩ @⁨Abhijeet CUBE⁩ @⁨Chithra Mam⁩ @⁨Himanshu Joshi⁩ & others .

Eric D. Green, M.D., Ph.D., is the director of the National Human Genome Research Institute (NHGRI) at the U.S. National Institutes of Health (NIH). Dr. Green’s relationship with the Institute began long before his appointment as director. He served as the Institute’s scientific director (2002 - 2009), chief of the NHGRI Genome Technology Branch (1996 - 2009) and founding director of the NIH Intramural Sequencing Center (1997 - 2009). Prior to that, he played an integral role in the Human Genome Project.Eric D. Green, M.D., Ph.D., is the director of the National Human Genome Research Institute (NHGRI) at the U.S. National Institutes of Health (NIH). Dr. Green’s relationship with the Institute began long before his appointment as director. He served as the Institute’s scientific director (2002 - 2009), chief of the NHGRI Genome Technology Branch (1996 - 2009) and founding director of the NIH Intramural Sequencing Center (1997 - 2009). Prior to that, he played an integral role in the Human Genome Project. - Google Search

Strawberry DNA Strawberry DNA Extraction

The average glass of wine contains about 11 percent to 13 percent alcohol, but bottles range from as little as 5.5 percent ABV to as much as around 20 percent ABV. 1 When tasting wine, you’ll notice alcohol comes through as heat in the back of your mouth or throat. Wine Alcohol Content Guide for Every Type of Wine.

Follow up discussion in Context to curriculum group:

[12/02, 10:07 pm] Khushi: detergents disrupt the cell membrane thus does cell lysis so that dna will come out of cell
[12/02, 10:34 pm] Himanshu Joshi:
What is this medium and how did you prepare this?
[12/02, 10:36 pm] Himanshu Joshi: How does detergent get to know that it has reached the cell membrane of a cell and disrupted only that?
What about Nuclear membrane? @⁨~Khushi⁩
[12/02, 10:37 pm] Khushi: so it interacts with the phospholipid memrane …
[12/02, 10:41 pm] Himanshu Joshi: How does this interaction take place, I mean when a soap solution comes in contact with cell membrane, what interaction of soap with phospholipid membrane take place and why?
[12/02, 10:42 pm] Khushi: so its hyphophobic and hydrophilic interaction between lipid membrane and detergent
[12/02, 10:44 pm] Himanshu Joshi: Okay, lipid membranes have hydrophilic and hydrophobic ends. Where does the detergent bind and why?

Khushi :Detergent monomers solubilize membrane proteins by partitioning into the membrane bilayer. With increasing amounts of detergents, membranes undergo various stages of solubilization. The initial stage is lysis or rupture of the membrane. https://r.search.yahoo.com/_ylt=AwrKBWxKUsplJ2UGaXDnHgx.;_ylu=Y29sbwMEcG9zAzEEdnRpZAMEc2VjA3Nj/RV=2/RE=1707786955/RO=10/RU=https%3A%2F%2Fwww.thermofisher.com%2Fus%2Fen%2Fhome%2Flife-science%2Fprotein-biology%2Fprotein-biology-learning-center%2Fprotein-biology-resource-library%2Fpierce-protein-methods%2Fdetergents-cell-lysis-protein-extraction.html%23%3A~%3Atext%3DDetergent%2520monomers%2520solubilize%2520membrane%2520proteins%2520by%2520partitioning%2520into%2Cstage%2520is%2520lysis%2520or%2520rupture%2520of%2520the%2520membrane./RK=2/RS=Os5PBvieDYrpuYMK37MlrPnG3gI-

[13/02, 6:38 am] Chithra Ma’am: Detergents can be denaturing or non-denaturing with respect to protein structure. Denaturing detergents can be anionic such as sodium dodecyl sulfate (SDS) or cationic such as ethyl trimethyl ammonium bromide. These detergents totally disrupt membranes and denature proteins by breaking protein-protein interactions.
https://www.google.com/search?q=does+detergent+denature+proteins&rlz=1CDGOYI_enIN944IN944&oq=does+detergent+denature+proteins&gs_lcrp=EgZjaHJvbWUyCQgAEEUYORiABDIICAEQABgWGB4yCAgCEAAYFhgeMgoIAxAAGA8YFhgeMggIBBAAGBYYHjIICAUQABgWGB4yCAgGEAAYFhgeMggIBxAAGBYYHjIICAgQABgWGB4yDQgJEAAYhgMYgAQYigXSAQkxNTk0OGowajeoAgCwAgDiAwQYASBf&hl=en-GB&sourceid=chrome-mobile&ie=UTF-8
[13/02, 6:39 am] Chithra Mam: So detergents can also denature proteins and create gaps in membranes, ultimately breaking it down.

Theertha · April 23, 2024, 5:42am

DNA EXTRACTION SETUP:CUBE HOMELAB PALAKKAD by Amritha , SN College Alathur.

Following are post made by Amritha in Context to curriculum WhatsApp group.

Date:22.4.24.
Time:12:56pm.
Experiment name:DNA extraction.
Name of things i used here
1.vim dish wash
2.1bowl.
3.Fruit name:watermelon
4.1 spoon.
5.2 glass.
6.hand sanitizer.
7.salt (i kept this salt in a steel bowl.
8.water(i take water in red bottle.
9.fliter.

First i put 2 pieces of watermelon

then i smash very well.
Date:22.4.24.
Time:12:56pm.
Place:palakkad.
Amritha.k sn college alathur.

I take one glass and filter and i kept this filter upper part of glass to take watermelon juice.
Date:22.4.24.
Time: 12:56pm.
Place:palakkad.
Amritha.k sn college alathur.

This is steel bowl i kept salt
Place:palakkad.
Date:22.4.24.
Time:12:56pm.
Amritha.k sn college alathur.

I add this water into steel bowl i kept salt.
Place :palakkad.
Date:22.4.24.
Time: 12:56pm.
Amritha.k sn college alathur.

This sanitizer i used here.
Ethyl alcohol IP. 80%v/v.
Glycerol IP. 1.45%v/v.
Hydrogen peroxide IP. 0.125%v/v.
Purified water. q.s.
Place:palakkad.
Date:22.4.24.
Time:12:57pm.
Amritha.k sn college alathur

Watermelon juice making.
Date:22.4.24.
Place:palakkad.
Time:12:56pm.
Amritha.k sn college alathur.

Adding water into a steel bowl.
Place:palakkad.
Date22.4.24.
Time:12:56pm.
Amritha.k sn college alathur.

Adding vim dish liquid into bowl.
Place:palakkad.
Date:22.4.24.
Time:12:56pm.
Amritha.k sn college alathur.

Adding extraction liquid into watermelon juice.
Place:palakkad.
Time:12:57pm.
Date:22.4.24.
Amritha.k sn college alathur.

Adding sanitizer into watermelon juice.
Place:palakkad.
Time:12:57pm.
Date:22.4.24.
Amritha.k sn college alathur.

Place:palakkad.
Time:12:58pm.
Date:22.4.24.
Amritha.k sn college alathur.

Place:palakkad.
Date:22.4.24.
Time:5:06pm.
Amritha.k sn college alathur.

Theertha · November 3, 2024, 2:15pm

Major questions addressed:

The primary focus of our discussion was the process of extracting DNA and how we can break the cell wall and cell membranes of onion cells to access the DNA. We drew a comparison between using hand wash during COVID times and the removal of dirt using detergent.

How can we break the cell wall and cell membranes of a cell?

Theertha · November 3, 2024, 5:42pm

S3E12 Cube chatShaala: Exploring new ways of Learning

In our college/institute laboratories, majority of us must’ve done DNA extraction and its further screening by extracting DNA from bacteria, etc and then running it on the AGE (agarose gel electrophoresis) equipment.
Can DNA be extracted at home?
Why? Why not!?
DNA is present in fruits/vegetables that we eat! We never bothered to look for it though😉.
How is the laboratory method of DNA extraction different from the HomeLab method?
I think the main requirements would be
i) a fruit/vegetable ii) alcohol (rum/whiskey? no! in COVID times, sanitiser with 70% alcohol is enough!) iii) detergent/soap iv) warm water v) salt.

In this slide from the whiteboard (almost a self-explanatory slide), we arrived at the conclusion that DNA can be extracted from things available at home, with ease! This was followed by a discussion on the importance of each requirement; mortar pestle (for grinding/making a paste), soap/detergent (to disrupt the cell membrane; as soap does to the COVID19), alcohol, etc.

Theertha · November 3, 2024, 5:45pm

"DNA Extraction Discussion:

Gaps and Misconceptions in Understanding Meiosis and Mitosis

Group photographs:2/11/2024

Theertha · November 4, 2024, 2:35pm

Summary made by @Enas in Context to curriculum whatsapp group

• How can DNA extraction be done at home and in a college lab?
• Is DNA and RNA soluble in water? If so, how does this solubility occur?
• How can we test for the presence of DNA?
• What is the purpose of each step in a DNA extraction protocol (e.g., crushing the fruit, adding detergent, adding salt, filtering, and adding alcohol)?
• Why does crushing a fruit (like a banana or watermelon pulp) break the cell wall but not the cell membrane?
• What kinds of contents or debris are present in the filtrate after filtering?
• How do we define the solubility of a compound?
• Are proteins soluble in water? Do water-soluble proteins contain non-polar amino acids?

Using hemoglobin as an example:
• Is hemoglobin soluble in water?
• Does hemoglobin contain non-polar amino acids, such as glycine, alanine, or valine?
What happens if a chain of amino acids containing both polar and non-polar amino acids is placed on the surface of water?

Theertha · November 5, 2024, 5:51pm

DNA EXTRACTION at CUBE Homelab by @Priti_Kanade

Context to curriculum whatsapp group discussions:

[05/11, 11:42 am] Priti CUBE: https://youtu.be/tTtw-ucKE6k?feature=shared
[05/11, 12:33 pm] Theertha: what is this DPA ? @⁨Priti CUBE⁩
[05/11, 6:59 pm] Arunan sir: Don’t you have some questions on the DNA extraction method? @⁨Theertha⁩ @⁨Amirtha CUBE⁩ @⁨Enas CUBE⁩ and others
[05/11, 7:14 pm] Priti CUBE: https://images.app.goo.gl/PXCbijGE8VcasZcy9

DPA+ DNA —> Blue color
This serves as an evidence that we extracted DNA
PFA
Chemical reaction
[05/11, 7:21 pm] Arunan sir : How did you manage to get DPA for the HomeLab? @⁨Priti CUBE⁩
[05/11, 7:29 pm] Priti CUBE: I had obtained from Ruia college
[05/11, 7:30 pm] Priti CUBE: Rest all materials are obtained at home

Theertha · November 5, 2024, 6:02pm

*Major questions addressed:5/11/2024

1)How to extract onion DNA?
2)Why was onion chosen?
3)Why is adding alcohol necessary to extract DNA?
4)Why is papaya juice added?

How does salt help in the extraction of DNA? Does it neutralize the charge of DNA? @⁨Enas CUBE⁩ @⁨Rechel CUBE⁩ and others

Why is SDS used in the standard DNA extraction method?

How does salt dissolves in water ?

Ionic bond or covalent bond,which is stronger ?

What happens when we add salt into water?

Reference shared:

Role of Salt in DNA extraction:

In general terms, salting out is the phenomenon observed when the solubility of a nonelectrolyte compound in water decreases with an increase in the concentration of a salt. https://www.sciencedirect.com/topics/chemistry/salting-out#:~:text=In%20general%20terms%2C%20salting%20out,need%20not%20concern%20us%20here.

What happens when we add salt into the water?
Water molecules and their interaction with salt https://www.usgs.gov/media/images/water-molecules-and-their-interaction-salt

Why Does Water Dissolve Sugar?Lesson 5.4: Why Does Water Dissolve Sugar? - American Chemical Society

Group photographs of participants taken at 9.50pm today :5/11/2024

Theertha · November 8, 2024, 11:33am

Objective: To extract the DNA from an onion.
Procedure:
1)I took one onion, crushed it (using a grinding mixy )and filtered it to get 4 spoons of onion extract.
Later ,added 2spoons of salt and stirred it for 5 minutes.
2)I added 1 tablespoon of washing powder,stirred it for 3 minutes.
3) added 1spoon of papaya juice,stirred it for 2 minutes.
4) added 10 ml of perfume.
Date:4/11/2024
Time:11am
Photo: Theertha

Procedure:DNA extraction from an onion.

Date:4/11/2024
Time:11am
Photo: Theertha

DNA extraction at Homelab, cherukkad Kozhikode
I can see the white clubbed part at the top as well as the bottom.
Is it the DNA? @⁨Amirtha CUBE⁩ and others
Date:4/11/2024
Time:11am
Photo: Theertha

[04/11, 12:21 pm] Amirtha CUBE: Please make video
[04/11, 12:24 pm] Amirtha CUBE: Take that white part theertha.
[04/11, 12:24 pm] Amirtha CUBE: Separate it
[04/11, 12:25 pm] Amirtha CUBE: Congratulations theertha
[04/11, 12:28 pm] Dinesh: Can you share stepwise procedure Dr Theertha?
[04/11, 1:33 pm] Dinesh: well done @⁨Theertha⁩
[04/11, 1:34 pm] Amirtha CUBE: Theertha please send photo of dna.

DNA Extraction at Homelab: Onion
Any tests to confirm this as DNA ?
Place: Cherukkad,Kozhikode
Date:4/11/2024
Time:11am
Photo: Theertha

[04/11, 2:30 pm] Theertha: DNA EXTRACTION: Onion
Requirements:
1 onion,Salt , Detergent,1 raw papaya and Perfume.
Place: Cherukkad,Kozhikode
Date:4/11/2024
Time:11am
Photo: Theertha
[04/11, 2:31 pm] Theertha: Objective: To extract the DNA from an onion.
Procedure:
I took one onion, crushed it (using a grinding mixy )and filtered it to get 4 spoons of onion extract.
Later ,added 2spoons of salt and stirred it for 5 minutes.
I added 1 spoon of washing powder,stirred it for 3 minutes.
I added 1spoon of papaya juice,stirred it for 2 minutes.
I added 10 ml of perfume.
Date:4/11/2024
Time:11am
Photo: Theertha
[04/11, 2:36 pm] Theertha: DNA EXTRACTION using Onion. Any confirmation test for DNA ? @⁨Amirtha CUBE⁩ @⁨Chithra Ma’am⁩ @⁨Rahul CUBE⁩ @⁨Enas CUBE⁩ and others
When this DNA is collected and purified, the result is a whitish, sticky substance that is somewhat translucent.DNA Is a Structure That Encodes Biological Information | Learn Science at Scitable.
Place: Cherukkad,Kozhikode
Date:4/11/2024
Time:11am
Photo: Theertha

[04/11, 2:47 pm] Amirtha CUBE: You add alcohol? theertha
[04/11, 2:50 pm] Amirtha CUBE: This is the white part you observe upper surface of liquid?theertha.
[04/11, 3:08 pm] Enas CUBE: Theertha, is it thread like when trying to take it out?
[04/11, 3:11 pm] Enas CUBE: Can we say that the precipitate is formed because of adding salt?
[04/11, 3:12 pm] Enas CUBE: What’s the percentage of alcohol present in the perfume Theertha?
[04/11, 3:12 pm] Dinesh: What’s precipitation?
Explain it to school students in understandable terms
[04/11, 6:15 pm] Arunan sir : Perfume? @⁨Theertha⁩
[04/11, 6:16 pm] Arunan sir : Salt? @⁨Theertha⁩
[04/11, 6:18 pm] +91 : Can we use sanitizer for alternative for ethanol?
[04/11, 6:21 pm] Arunan sir : Which white part?
Why one settled in the bottom and other floating? @⁨Amirtha CUBE⁩ @⁨Theertha⁩
[04/11, 6:21 pm] Arunan sir : Great initiative. Congratulations @⁨Theertha⁩
[04/11, 6:26 pm] Arunan sir : Why salt in the second step? @⁨Theertha⁩
What are you expecting to happen?
[04/11, 6:27 pm] Arunan sir : What does washing powder do in DNA extraction from Onion extract? @⁨Theertha⁩
[04/11, 6:28 pm] Subhashree: Why have u added papaya juice in this procedure ? What it’s significance? Can u pls share ?
[04/11, 6:29 pm] Arunan sir : Great initiative @⁨Theertha⁩
Why papaya juice?

Are you extracting DNA from Papaya too?
[04/11, 6:29 pm] +Batul: Yes, I have the same question @⁨Theertha⁩ when you are extracting DNA from onion why have you added papaya juice
[04/11, 6:30 pm] Arunan sir : Perfume? @⁨Theertha⁩
Which perfume?
[05/11, 11:42 am] Priti CUBE: https://youtu.be/tTtw-ucKE6k?feature=shared
[05/11, 12:33 pm] Theertha: what is this DPA ? @⁨Priti CUBE⁩
[05/11, 1:08 pm] Theertha: Sorry for referring to it as perfume. I need alcohol to precipitate the DNA, so I used perfume as an alternative since it contains alcohol.
Composition:
Alcohol Denat, Aqua, Parfum. PEG:40 Hydrogenated Casior Oil Amyl Cinnama! Benzy! Alcohol. Ber-zy Salicylate Citral, Citronelloi Coumarin, Geraniol, Hydroxycitronellal, Isoeugenol, Limonene, Linalool
Buy Brut Edt Original (Plexi Box) Silver Edition, 100ml Online at Low Prices in India - Amazon.in
[05/11, 1:09 pm] Theertha: Here, the salt is sodium chloride (NaCl). Sorry for referring it to simply as salt.As sodium in the salt can bound with phosphate in the DNA and create a attraction between them.
Little amount of salt also can keep the protein dissolved in water.
[05/11, 1:11 pm] Theertha: Yes ,i do think so.Salt cause the protein to precipitate here.
[05/11, 1:12 pm] Batul: Why did you add papaya juice?
[05/11, 1:13 pm] Theertha: I read that t he cell wall and cell membranes can be destroyed by hydrolysis of water. In the next step, I added salt since the cell wall and membranes were already destroyed. Adding detergent later will break apart the lipid bilayer.
This video also follow the same way:

Hydrolysis of water with grinding can break the cell wall and cell membranes ,both glycosidic bond and lipid bilayer? @⁨Amirtha CUBE⁩ @⁨~Khushi @⁨Chithra Ma’am⁩ and others
[05/11, 1:14 pm] Theertha: I found that meat tenderizer used for DNA extraction,so instead i used the papaya juice.
Papaya contains , papain which can break the proteins
@⁨~Batul Pipewala⁩ @⁨~subhasree vinodhkumar⁩
[05/11, 1:15 pm] Theertha: Washing powder is made up of Sodium /pottasium salt of fattyacids which can forms a micelles as hydrophobic & hydrophilic part of both washing powder and lipid layer will form a micelles will cause the breaking of cell membranes
[05/11, 1:16 pm] Khushi: yes hard grinding with mortar n pestl or mixer grinding can break the cell walls easily …this is a common technique to break cell wall
[05/11, 1:17 pm] Theertha: Major goof-ups I made: Looking for the critical analysis and suggestions for the next setup @⁨~Batul Pipewala⁩ @⁨~Khushi⁩ @⁨~subhasree vinodhkumar⁩ @⁨Enas CUBE⁩ @⁨Kiran CUBE⁩ @⁨Arunan sir and others

1)Added too much salt, which caused the precipitation of the protein!
2)The perfume I used did not indicate the amount of alcohol it contain, possibly a lower amount of alcohol wouldn’t have precipitated the DNA.
3)Using papaya juice and onion juice did not align with my objective, which was to isolate onion DNA.
[05/11, 1:20 pm] Khushi: u can use saintiser which has 70 percent alchohol …
[05/11, 1:21 pm] Khushi: onion is commonly used to isolate DNA
[05/11, 1:29 pm] Vikram crishna: Please check with a chemist shop, I think it is possible to buy small quantities of methyl alcohol (50-100ml), maybe under the name methylated spirit.
[05/11, 1:35 pm] +Sandhir: Spirit is another choice
[05/11, 1:36 pm] sandhir: It would have additives which may not affect the results
[05/11, 6:59 pm] Arunan sir : Don’t you have some questions on the DNA extraction method? @⁨Theertha⁩ @⁨Amirtha CUBE⁩ @⁨Enas CUBE⁩ and others
[05/11, 7:04 pm] Arunan sir : Let’s discuss this and get more clarity during Chatshaala. @⁨Theertha⁩ @⁨Enas CUBE⁩ @⁨~Batul Pipewala⁩ @⁨Amirtha CUBE⁩ @⁨Rahul CUBE⁩ and others.
[05/11, 7:05 pm] Priti CUBE: What’s the use of papaya juice
[05/11, 7:06 pm] Priti CUBE: What were the contents of the perfume
[05/11, 7:06 pm] Arunan sir : Looks very basic, like grinding for Dosa dough. Please join Chatshaala @⁨~Khushi
[05/11, 7:09 pm] Arunan sir : Let’s discuss with all others during Chatshaala. @⁨Theertha⁩ @⁨~Mohammad Yousuf Ganaie⁩ @⁨Enas CUBE⁩ @⁨~Batul Pipewala⁩ @⁨Priti CUBE⁩ and other Cubists.
[05/11, 7:14 pm] Priti CUBE: https://images.app.goo.gl/PXCbijGE8VcasZcy9

DPA+ DNA —> Blue color
This serves as an evidence that we extracted DNA
PFA
Chemical reaction
[05/11, 7:17 pm] Arunan sir : What is the role of alcohol in extracting DNA? @⁨Theertha⁩ @⁨~Vickram Crishna⁩ @⁨Enas CUBE⁩ @⁨~Mohammad Yousuf Ganaie⁩ @⁨Priti CUBE⁩ @⁨~Khushi ⁩ and others.
[05/11, 7:18 pm] Arunan sir : How does spirit help in extraction of DNA?

Where do we get spirit from? @⁨~Rajat Sandhir⁩
[05/11, 7:21 pm] Arunan sir : How did you manage to get DPA for the HomeLab? @⁨Priti CUBE⁩
[05/11, 7:25 pm] Arunan sir : Please join Chatshaala today for continuing the exciting discussion happening in CUBE groups on Extraction of DNA in HomeLabs
and other exciting questions, for better clarity.

https://webinar.hbcse.tifr.res.in/b/kir-qwu-8bc-2ur
[05/11, 10:46 pm] Vikram crishna
: One assumes the purpose of isolating the DNA of the onion is not to later consume the onion DNA, so the preferred solvent will be methyl
[05/11, 10:55 pm] Priti CUBE: Cubists have extracted dnA with all basic ingredients
And yes using perfume as well
[05/11, 10:56 pm] Priti CUBE: What is the purpose of using alcohol in dna extraction?
Is it toxicity?
[05/11, 11:16 pm] Vikram crishna
: I am learning about DNA extraction for the first time, only from the discussions and trials here.

It seems to me that to do an experiment well, it is best to use the most suitable materials. In this case, the best solvent, methyl alcohol, for this application.
[06/11, 11:49 am] Vikram crishna
: My point is, I don’t know if those contaminants will change the chemical form of the isolated DNA.

So, I take your point, learning how to do the basic extraction is a good thing.

The next step will be to work towards useful extraction, a pure form of the DNA that can be kept for the next stage of studies.

For my learning, please outline what kind of studies are possible in a homelab, and further, what kind of discussion we can have that opens up new possibilities.

I should explain first that it has never occurred to me that this complex molecule, the stuff of Nobel prizes, could even be accessible to a homelab.
[06/11, 11:57 am] Arunan sir : That’s what Cubists like @⁨Amirtha CUBE⁩ @⁨Theertha⁩ etc. are venturing.
The most important aspect is that they are open and not secretive while doing this experimental project and are open to collaboration and criticisms .
The latter is rare in Indian Science Education and Research.
Please join them in this and other HomeLab activities.
HomeLab has a moto that nothing is alien to any one, as, if there is a will, there is a way.
Let’s all join in this revolution.
[06/11, 12:04 pm] Vikram crishna: Let’s be very fair, openness is not a hallmark of science anywhere, particularly not in educational ‘institutions’. There are different reasons, in different countries and social environments, but the situation ends up the same, preserving the status quo for the already privileged.

May I suggest that the short videos here be uploaded with comments/captions to public platforms.

Most of these are, it is true, captured by vested interests, either US or Chinese billionaires, but there are a few others equally accessible, even if not all of them have published separate channels for ‘short’ lessons.
[06/11, 3:33 pm] Theertha: Alcohol can precipitate the dissolved DNA in the water .
Reference:Ionic salts and DNA both readily dissolve in water but not so easily in ethanol or isopropanol because they both require a polar solvent such as water to dissociate. Adding ethanol or isopropanol to a salty solution, therefore, causes the DNA to pellet and precipitate out of solution.
https://onlinelibrary.wiley.com/doi/10.1002/nano.202100152#:~:text=Ionic%20salts%20and%20DNA%20both,and%20precipitate%20out%20of%20solution.
[06/11, 3:34 pm] Theertha: salting out can cause protein to precipitate.

Reference:
Salting-out is another simple DNA isolation method. In this procedure, cells or tissues are first lysed and treated with proteinase K and RNase. The use of saturated NaCl results in protein precipitations.
Qualification Study of Two Genomic DNA Extraction Methods in Different Clinical Samples - PMC.
[06/11, 3:36 pm] Theertha: Can we use sanitizer as an alternative of alcohol ,to precipitate the DNA? @⁨~Khushi ⁩ @⁨~Vickram Crishna⁩
[06/11, 3:37 pm] Theertha: Where do we get the spirit? Any homelab materials have spirit?
[06/11, 3:38 pm] Theertha: SDS is doing the same way as detergent doing ,it helps to break the cell membranes and nuclear membranes.
Reference:
Strong ionic detergent SDS helps to solubilize membrane proteins and lipids, thereby causing the cell to lyse and release its contents.
Comparative proteoinformatics revealed the essentials of SDS impact on HaCaT keratinocytes - PMC.

Theertha · November 8, 2024, 3:22pm

Major goof-ups I made:

1)Added too much salt, which caused the precipitation of the protein!
2)The perfume I used did not indicate the amount of alcohol it contain, possibly a lower amount of alcohol wouldn’t have precipitated the DNA.
3)Using papaya juice and onion juice did not align with my objective, which was to isolate onion DNA.

Biochemistry of DNA Extraction!

1)Breaking the cells (cell disruption) and release of the DNA can be accomplished using appropriate methods including grinding the living tissues.

(2)Removing membrane lipids by adding a detergent.

(3)Cellular proteins including histone that bound to the DNA can be removed by extracting with a phenol-chloroform mixture or precipitating with sodium or ammonium acetate or adding a protease.

(4)Precipitating the DNA with alcohol (usually isopropanol or ethanol). Since DNA is insoluble in these alcohols, it will aggregate together, giving a pellet upon centrifugation. @⁨Amirtha CUBE⁩ @⁨Enas CUBE⁩ @⁨Rechel CUBE⁩ and others.
https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/dna-extraction
Biochemistry of DNA Extraction!

1)Breaking the cells (cell disruption) and release of the DNA can be accomplished using appropriate methods including grinding the living tissues.

(2)Removing membrane lipids by adding a detergent.

(3)Cellular proteins including histone that bound to the DNA can be removed by extracting with a phenol-chloroform mixture or precipitating with sodium or ammonium acetate or adding a protease.

(4)Precipitating the DNA with alcohol (usually isopropanol or ethanol). Since DNA is insoluble in these alcohols, it will aggregate together, giving a pellet upon centrifugation. @⁨Amirtha CUBE⁩ @⁨Enas CUBE⁩ @⁨Rechel CUBE⁩ and others.
https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/dna-extraction

[06/11, 4:57 pm] Arunan sir : Do you agree with this explanation from the reference sent by @⁨Theertha⁩
@⁨~Mohammad Yousuf Ganaie⁩ @⁨Kiran CUBE⁩ @⁨Rechel CUBE⁩ @⁨Enas CUBE⁩ and others.

“According to Rice (2018), the method involves three necessary steps, namely, lysed, precipitation, and purification. The first step is lysed to break down the cells so the DNA in the nucleus will be released after the cell membrane is disrupted. The second step is precipitation. To get pure DNA, the DNA will be separated from proteins and other cells component by degrading the DNA and protein with ethanol or isopropanol.”
https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/dna-extraction#:~:text=According%20to%20Rice,or%20isopropanol.

Theertha · November 8, 2024, 4:15pm

Second attempt to isolate DNA from onion

Improvement included:
1)Added salt after adding detergent.
2)Used sanitizer instead of Perfume,as the % if alcohol content not mentioned in the perfume bottle.
Requirements:
1 onion
Washing powder (detergent)
Sodium chloride (salt)
Sanitizer (contains alcohol)
Procedure:
1)I took one onion and extracted the juice by grinding it in a mixy (mechanical breakdown of cell walls and cell/nuclear membranes).
2)I added a 1 spoon of washing powder (detergent) to break down the lipid bilayer of the cell /nuclear membranes and Stirred for 3min
3)added 2 spoons of salt (sodium chloride, table salt) to precipitate the proteins (salting out).Stirred for 2min.
4)added sanitizer, which contains alcohol, to precipitate the DNA.

Result:
The white clubbed part observed on the surface may be of DNA
F* urther step required:*
The identification or confirmation test for DNA.
Date:8/11/2024
Time:12pm
Place: Cherukkad Kozhikode
Theertha

[08/11, 7:00 pm] Chithra Ma’am: What about the precipitated proteins in the clubbed part.
You missed an important step - filter the precipitate before adding alcohol/sanitiser.

[08/11, 9:27 pm] Theertha: Yes🙈.I am also concerned about why I am not getting a white thread/all DNA clubbed when I try to pull out the topmost white part! Is it not DNA @⁨Chithra Ma’am⁩
[08/11, 9:32 pm] Himanshu Joshi: Filtering is an important step in DNA isolation as proteins also get along with DNA, while adding alcohol.
Or
Another reason could be salting out that happens at high concentration helping in getting rid of the proteins.
Focus on these two steps and you would get success!
All the best @⁨Theertha⁩