We're going to go through the steps of what it takes to extract a sample of your dna.The first thing we need to do is actually get a sample of our cells so that we can extract the dna from our cells.So we're going to use cheek cells to get a large amount of dna that we can actually extract from them.The first thing we need to do is actually get a sample of our cheek cells, so i'm going to actually gargle my mouth with this water and then that will move around enough epithelial.
Cells from my cheeks to be able to get them out of my mouth.So in that water then we should have enough epithelial cells for our extraction.So i'm going to pour the water back into my tube.The next thing i'm going to do is i'm going to add some lysis buffer.In the lysis buffer there is a detergent and the detergent works just like soap does or shampoo or a dish detergent that we would use in our dishwasher.It's going to break down the components that are sitting here in my cell extraction.So i'm.
Going to add 2 ml of lysis buffer.And the detergent is going to start breaking down this extraction.So what the detergent will do is that it'll break open two membranes.It will break open the cell membrane on the outside of the cell and it'll also break open the nuclear membrane which of course is where our dna is housed.So i'm going to gently invert this four or five times.And what this is going to do is it's basically going to get the detergent in contact with everything in the solution.
And start breaking open those membranes.We don't want to shake it too hard because that could actually break our dna into small pieces and then we wouldn't be able to visualize it.So just a gentle mix is sufficient.The next thing we're going to do is we're going to add protease.Protease is an enzyme that breaks down protein so that will start breaking any large proteins that are still held together inside our solution.So i'm going to add 20 l of protease into my extraction.And again i'm going to gently invert my extraction four or five times to thoroughly mix the protease.
Next thing i'm going to do is i'm going to incubate my cell extraction at 50c.That is the optimal temperature for protease activity and it's going to incubate there for 10 minutes.So our incubation with our protease is over.So we're going to take our cell extraction out of the 50 water bath.And the next thing we're going to do is add cold alcohol.And the cold alcohol is going to facilitate the extraction of dna out of the solution.The alcohol is on ice.And i'm going to add the alcohol at an angle and slowly add it.
In to our extraction so that the alcohol runs down the side of the tube and i'm doing this slowly to create two layers the aqueous layer and the non aqueous layer.The combination of the alcohol plus the lysis buffer will allow the dna to come out of solution so that we can visualize it.So we'll leave that extraction to sit for 5 minutes undisturbed.So we've allowed our extraction to sit with the alcohol for 5 minutes.What i'm going to do now is i'm going to slowly.
Invert my tube so that the alcohol layer mixes with the aqueous layer.As i'm mixing, the dna will come out of solution and will form an aggregate and we'll see that aggregate as white threads at the interface where the alcohol and aqueous solution meet.So when we stop mixing it, we can see the white strands in our extraction solution and those long thin white strands that is your dna.So once we have the dna extracted, there are a few more experiments we can perform.We can take the dna and cut it into smaller fragments using restriction enzymes.We can also use.
361L AcidBase Extraction 5
So, i am going to separate a mixture that contains an organic acid, an organic base, and a neutral compound using acidbase extraction.Specifically, i will dissolve a mixture that contains benzoic acid, pbromoaniline, and phenanthrene in dichloromethane, and then throw it into a separatory funnel.Then i will add a solution of hydrochloric acid, which will convert the pbromoaniline into it's hydrochloride salt, which can then be removed in the aqueous layer.Once that's removed, i will throw in a solution of sodium hydroxide, which will convert the benzoic acid into a sodium salt, which can then also.
Be removed in the aqueous layer.Once the three compounds are separated, i will convert them all back to their neutral solids.Benzoic acid is corrosive.Phenanthrene is an irritant.Pbromoaniline is an irritant.Dichloromethane is toxic.Here i have a one to one to one by mass mixture of benzoic acid, phenanthrene, and pbromoaniline.I going to dissolve the mixture in ten milliliters of dcm or dichloromethane.I am going to add the solution to a separator funnel.Next, i am going to add approximately five milliliters of three molar hydrochloric acid.Two layers are noticeable.I am going to cap the sep.
Funnel and shake it to mix the two layers so the pbromoaniline that is in the dcm layer will come in contact with the hcl and cause protonation.The pbromoaniline hcl salt will then migrate to the aqueous layer.I will vent it so the pressure does not build up too much.Shake some more, vent again, shake one more time, and vent one more time.To be safe, i am going to add a few more drops of dcm and watch where they go so i know which layer is the dcm layer and which layer is the aqueous layer.You can see the drops fall.
To the bottom.This makes sense because dcm has a greater density than water and is usually the bottom layer.So i am going to label this bottom flask with an n , which will be the neutral, organic, dichloromethane layer.And now i will just drain off the bottom layer being careful not to get any of the top layer.I am going to label a second flask a.E., which will be the acidic extract or aqueous layer containing the hcl and the pbromoaniline salt.I will drain the solution into the flask, close the sep funnel, and add back the neutral.
Dcm layer so i can extract it one more time to make sure i get all the pbromoaniline out of it.I will add approximately another five milliliters of three molar hcl, cap the sep funnel, shake it, vent it, shake it, vent it, drain off the bottom, neutral dcm layer, and then collect the top aqueous acidic extract layer.I will pour back the neutral layer that should no longer contain pbromoaniline, but still contains benzoic acid and phenanthrene.To remove the benzoic acid, i will add approximately five milliliters of three molar sodium hydroxide.
I will shake the sep funnel to facilitate the deprotonation of benzoic acid by sodium hydroxide, forming ionic sodium benzoate which will migrate to the aqueous layer.And as before, just to be safe, i will add a few drops of dcm.The dcm as expected is the bottom layer.I will label a new flask b.E.For the base extracted aqueous layer.First though, i will drain off the bottom, neutral dcm layer, and then collect the base extracted aqueous layer.I will close the sep funnel and pour back in the neutral dcm layer, then extract.
It one more time with approximately five milliliters of the three molar sodium hydroxide solution.You can sort of see two layers on the sides of the sep funnel, but not in the middle.I'm going to add some more three molar sodium hydroxide to see if i can get the layers to be more noticeable.Now i will drain off the bottom, neutral dcm layer and then collect the second portion of the base extracted aqueous layer.So now all three compounds should be separated.The acidic extract should have the pbromoaniline.
Hydrochloride salt.The basic extract should have the sodium benzoate.And the neutral dcm layer should still have the phenanthrene.So now i need to turn them all back into the neutral solids they once were.I'm going to add anhydrous sodium sulfate to the neutral dcm layer to absorb any water that may be present.You can tell enough has been added when you can swirl it around and its not all completely clumped together.I will set this aside for now.Next, i will add six molar hcl solution to the basic extract, which will protonate the sodium benzoate,.
Converting it back to the benzoic acid.I will monitor the ph of the solution to ensure the protonation occurs.Right now the basic extract has a ph of about 10 and the six molar hcl has a ph of about 2.I will know the protonation is complete when the basic extract also has a ph of 2.I will grab some ice to cool the basic extract before the addition of the hcl solution since some sodium hydroxide is also present and to minimize the solubility of the benzoic acid.For similar reasons, i will also start cooling the acid extract and some.
Distilled water to help during transfers.I will add some of the hcl solution, and then take the flask out of the icewater bath so it can be seen easier.Formation of benzoic acid as a white precipitate is noticeable.After adding approximately five milliliters of the hcl solution, i will now test the ph, which is the same as the hcl solution and confirmation that all of the sodium benzoate has been converted to benzoic acid.I will collect the solid using vacuum filtration.I will wet the filter and attempt to transfer.
As much of the benzoic acid as possible.I will then rinse the flask a few more times and then leave the solid to dry on the vacuum for awhile.When it's dry, i will collect it on a watch glass and record its mass.Similar steps are used to convert the pbromoaniline hcl salt back to the free base form.In this case, six molar sodium hydroxide is used instead of the six molar hcl and when addition of the sodium hydroxide solution is complete, the acidic extract should have a ph of about 10.
After at least ten minutes of drying with sodium sulfate, the neutral dcm extract, which should still contain phenanthrene, can be transferred to a beaker, leaving behind the solid sodium sulfate and the excess water it absorbed.The dcm may uncontrollably drip out of the pipe.To prevent this from happening during transfer, it can be sucked up a few times to equilibrate the inside of the pipet before attempting to transfer it.To prevent any of the sodium sulfate from entering the pipet, positive pressure can be applied to the pipet bulb until it reaches the very bottom of the flask.To ensure i did not leave behind.
Any of the phenanthrene, i will rinse the flask with a few more milliliters of additional dcm and then transfer that as well.Next, i will evaporate the dcm solution on a hotplate in a sand bath.Instead of a boiling chip, i will add a wood applicator stick.Dcm has a low boiling point at forty degrees celsius.To prevent burning the solution when there is only a little bit of it left, i will make sure it is no longer in direct contact with the sand.I will also let the very last of it evaporate on its own at room temperature.
And increase its surface area by rolling it around a few times in a beaker.I can then scrape it from the beaker to a watch glass and when its completely dry, record its mass.I started with 520 milligrams of the mixture.I ended up with 100 milligrams of pbromoaniline, 107 milligrams of benzoic acid, and 145 milligrams of phenanthrene.So the percent recovery was 68 percent.This value makes sense because i lost some of it during transfers.The highest recovery for phenanthrene may be due to its greater tendency to always stay within the.
Acid Base Extraction Demonstrated By Mark Niemczyk PhD
Acid base extraction demonstrated by mark niemczyk phd,Acid base extraction demonstrated by mark niemczyk phd. 361l acidbase extraction 5,A mixture containing pbromoaniline benzoic acid and phenanthrene is separated using acidbase extraction closed captions available chemistry lab at. Acid alkaline diet foods alkaline foods,Shopliferegenerator reboundliferegenerator the foundation of all alkaline diets is to of course alkalize the body by consuming.
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Alkaline lysis miniprep,For more information visitbioradytnucleicacidpurification this tutorial demonstrates the alkaline lysis method of purifying plasmid dna from a.
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DNA Extraction By Phenol Chloroform
Dna extraction by phenol chloroform,Abnova this method relies on phase separation by centrifugation of a mix of the aqueous sample and a solution containing phenol and. Plasmid isolation for bacteria,Reagents and solutions alkaline solution i 50 mm glucose 25 mm tris hcl ph 80 10 mm edta ph 80 10 mgml lysozyme alkaline. Ellyn mulcahy dna extraction,Professor ellyn mulcahy demonstrates dna extraction at johnson county community collegejcccedu.
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Intervene Herbal Testimonials The Cure For Acidity Acid Vs Alkaline
Intervene herbal testimonials the cure for acidity acid vs alkaline,Individuals who started out skeptical have been amazed at the power and results of this nano blend of tree oils and plant extracts this health supplement is. Grapefruit has quinine propertiesextreme sicknesslupusleg crampscancer,Fatigue grapefruits are also beneficial in the treatment of fatigue so it can help you to dispel your general tiredness caused from routine or boring work. Health benefits of turmeric,Draxe in todays tutorial i want to share with you the incredible benefits of turmeric there are over 6000 al studies proving turmeric to be the 1.
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