DNA Extraction Protocol - Part 1 

Thanks for the question. Heat incubation activates the enzyme that will break down the cells and nuclei to expose the DNA.

thanks :)

@@jacksonlaboratory For this why don't we use strong detergents for cell lysis? and then enzymes for for degrading nuclear membrane so that DNA gets exposed?

I think to sterilize it good question

@@ektagupta7504 STRONG detergents are used for cells with cell walls (and hard ones of course). This experiment's sample were saliva spat by students which is linked with animal cells. Animal cells do not have a cell wall only a cell membrane, hence strong detergent is not advised, scientifically to be used since it might quickly disrupt the cell membrane and denature the very genomic DNA we want to extract. Hence enzymes are mostly preferred...

Lol, yes I am 😂

All of these "proprietary" solutions involved with DNA are usually salt solutions. They make them proprietary to keep that a secret because people do not want to pay 50 bucks for an ounce of salt water.

For accuracy

Hi Trasvi. The purifying solution is a proprietary solution from DNAGenotek that precipitates cell debris as part of their saliva extraction kit.

Ignore my statement below; it is an incorrect conjecture that would need to occur later in the process and only if they were planning on performing gene splicing. They haven't explained what they plan to do after extracting the DNA. The sample is mixed with the purifying solution AFTER heating the sample. I believe that they are using it to break down and dissolve the cell walls. Then they begin the annealing process (the ice). From there they use the centrifuge to separate the lighter material (dna) from the heavier material (leftover dissolved larger scale cellular detritus - the white pellets). ---- Look up 'Genetic Engineering - Isolating a gene'. Now I am not an expert in this subject -- but if I had to guess; the 'purifying solution' contains either one or many types of 'Restriction Endonucleases' that are used to split the DNA into discrete pieces prior to putting the DNA into their gene sequencer.

Salt water

Same question

It all depends on the protocol different extraction kits have slightly different protocol but most are similar

Hi, write me to tomas.hluska at upol.cz I can translate our "home-made" method for you (I think it's from Sambrook anyway) that we used in our lab course.

nothing happens, surface tension keeps the water at the bottom of the tube

In the case you're describing the pellet has the cells in it. In this case, the DNA is contained in the supernatant while impurities (protein, cell wall components, etc.) are contained in the pellet because the purifying solution added to the mixture (that made it cloudy) denatures the cells. The DNA will not precipitate out into a pellet unless you add specific reagents which usually come in another kit. Whether you keep the supernatant or not largely relies on your end goal, and what reagents and procedures you are using.

That depends which part is important for you. In case of bacteria storage, you pellet the bacteria which you are interested in and thus proceed with the pellet. In case of DNA extraction, you actually use both (in two separate centrifugation steps; unless you use binding columns). First, you precipitate and sediment the proteins which you want to get rid of, while your DNA is in the solution. Hence you work with the supernatant. In the second step, you precipitate your DNA with alcohol and hence you proceed with the pellet.

If you are dispensing the pipette yes. It’s recommended to have it at a 45 degree angle and against the wall of the tube you are dispensing into. It will help “pull” all the volume out.

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You aren't kidding--a country song for a high-tech topic.

It is DNA from mouse tissue.

@@jacksonlaboratory from mouse tissue? Didn't you say that students spit into the tubes?

@ The students did spit in the tubes, but they were instructed to chew the mice first.

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ru-vid.com/video/%D0%B2%D0%B8%D0%B4%D0%B5%D0%BE-QYpX94prb0A.html Protocol 2 - PCR

the detergent disrupts the cell and nuclear membranes of each cell to release the DNA. It does this by dissolving lipids and proteins that hold the membranes together.

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The lysis step. SDS is what you would call the soap. This is one of the first steps, along with a enzyme that breaks down proteins. Then your isopropyl/etoh (alcohol), which precipitates out the DNA since it's not soluble in alcohol.

E. coli has tough cell wall, so I don't think so. You need alkaline lysis for bacteria. qPCR requires clean DNA what should be provided by any kit method.

@ or heat... Bacteria break apart easily, 10 min at 95C

@@AndrewLyon23 we used heat for direct genotyping bacteria by PCR, but does qPCR work as well?

Good practice!!

Short answer - yes! www.ncbi.nlm.nih.gov/pubmed/18791146

For dna extraction from hair, is it the same method as this?

Ok

For accuracy

cold reduces solubility of impurities.

why do you want to extract DNA from dead?

A little bit too suspicious buddy

@@avolatube_8253 ... forensics

are you asking for a friend?

I don't think so. Plants have cell wall, which need to be broken. Often various protocols using CTAB are used.

most likely.

If blood observed on nasal swab kit during taking sample from nasal, it will be cause of covid-19