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Totipotent Stem Cell From Molecular Components

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caters
#1
Feb8-14, 12:40 PM
P: 38
I am making a human stem cell from its molecular components(just pretending to do it right now but might be able to do later in life).

I have extracted my DNA and formed a liposome.

I have the DNA replicating in an incubator at 98.6 F, 5% CO2, and 95% humidity.

The temperature is because it is human.
The CO2 is so that it has a stable pH.
The humidity is so it does not get dehydrated.


I have calculated how much DNA I will have in a week. Here are my calculations:

8 2/3 hours for replication(10 base pairs per second because a process always goes as slow as the slowest enzyme)

Doesn't matter how many replication origins there are.

8 2/3 hours per round = 26/3

24 hours per day divided by 26/3 hours per round is the same as 24/1 * 3/26

That gives you 72/26 = 36/13 = 2 10/13 rounds per day

That is the number of rounds of replication for 1 day.

Multiply by 7 to get 252/13 = 19 5/13 rounds per week

Round that to 19


Than remember that DNA replicates in powers of 2

2^19 = 524288 total chromosomes from 1.

Is that only approximate?

Also I am planning to do a microarray analysis of differentiated cells and types of stem cells against my totipotent stem cells(ES cells without hurting an embryo) and I don't want to run into epigenetic problems so should I have a biopsy done of all my tissues and then be treated for any health problems that result such as ulcers and heart problems? I mean if thats the case I will have my own tissue to separate into cells and won't run into epigenetic problems. If not than I might run into epigenetic problems. Like one person's cardiac muscle might have a different amount of protein than mine and thats a problem.
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Ryan_m_b
#2
Feb11-14, 06:29 PM
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I'm sorry but I really can't follow what you're trying to propose here. What is driving DNA replication? DNA within a liposome is not enough. How would any of this lead to a stem cell, let alone many different types of cell?

If you're interested in the topic of making stem cells I suggest reading up on iPSCs.
caters
#3
Feb11-14, 06:42 PM
P: 38
The DNA is being replicated by enzymes in a test tube inside an incubator. The chromosomes are separated before then. I separate 1 pair of chromosomes from each test tube for putting inside the nucleus. I do PCR with the others to have the gene for a particular protein replicated. I then have a test tube in which mRNA and then protein is formed. Than I separate the newly formed protein from the rest of the stuff in the test tube by centrifugation. I then use microtools to put the protein wherever it needs to go.

caters
#4
Feb11-14, 06:43 PM
P: 38
Totipotent Stem Cell From Molecular Components

and IPS cells cannot become ANY cell whereas my totipotent cell will. Plus you don't have to hurt an embryo to get totipotent stem cells if you do it this way.
gravenewworld
#5
Feb11-14, 08:08 PM
P: 1,405
This thread doesn't make any sense and should be locked/deleted.

There are a huge number of things wrong with your ideas and you should go back and re-read a bio101 text on how a protein is made from DNA which includes things like control mechanisms to make sure proteins are folded correctly (which you don't even have in your soup). You made a protein from DNA, so what? Is it properly folded and post-translationally modified correctly to make it into even a biologically active/relevant/properly folded form? Post-translational modification such as glycosylation, sulfation, acetylation, phosphorylation, ubiquitination, isoprenylation, GPI anchoring of a cell surface protein, etc. etc. etc., which are all critical for a properly functioning cell and proteins, can not be controlled with a template like proteins/DNA.


Sorry, you're out of luck. We won't even be able to do this in the next 250 years I'm guessing.
caters
#6
Feb11-14, 09:14 PM
P: 38
with the advance in nanotechnology I would say sooner than 250 years for the tools. And with how fast we are advancing in cellular biology I would say sooner than 250 years for that.
Ryan_m_b
#7
Feb12-14, 07:27 AM
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Is English your first language Caters? It's quite difficult to follow what you are talking about exactly.

Quote Quote by caters View Post
The DNA is being replicated by enzymes in a test tube inside an incubator. The chromosomes are separated before then. I separate 1 pair of chromosomes from each test tube for putting inside the nucleus.
Let me see if I've got this straight: you're proposing a system in which you have test tubes containing a reaction to replicate DNA. It's not just replicating the sequence but somehow building a functional chromosome with all the support proteins needed (histones for example). How do you propose this is possible? What is going on inside these tubes to achieve this? Furthermore you're proposing that each tube makes a different chromosome and you then are going to combine these into a, presumably empty, cell nucleus right?

Assuming I'm correct that this is what you mean what is the point if all this? At the end you're just going to have a cell which you could have just got by culturing the original one you harvested the DNA from. And where does totipotency come in here? Whether or not a cell is totipotent, pluripotent and what kind of cell it is isn't governed by the DNA sequence. Every cell has the same genotype, it's the complement of what genes are on and what are off that determine cell behaviour (simplistically, the microenvironment obviously plays a role).

Quote Quote by caters View Post
I do PCR with the others to have the gene for a particular protein replicated. I then have a test tube in which mRNA and then protein is formed. Than I separate the newly formed protein from the rest of the stuff in the test tube by centrifugation. I then use microtools to put the protein wherever it needs to go.
So now you seem to be mixing in protein manufacturing but I don't understand what you mean here. In a separate system you want to PCR for a particular gene, then in a cell free manner produce large quantities of that protein. Whilst there are methods of doing this the most established is recombinant DNA technology. Regardless I don't see what this is meant to achieve. And what is a microtool?

So at the moment all you seem to have is a complicated way of replicating a cell without any real outlining of how you're going to do it (other than putting DNA in an empty nucleus, presumably with the cytosol and organelles intact) and an unrelated system building protein. What does any of this have to do with manufacturing totipotent cells?

Quote Quote by caters View Post
and IPS cells cannot become ANY cell whereas my totipotent cell will. Plus you don't have to hurt an embryo to get totipotent stem cells if you do it this way.
You need to check your terminology. Pluripotent cells are defined as cells that can differentiate into any cell of the three germ layers (I.e. Any cell in an organism). A totipotent cell is the same but can also make placental cells which are not important post-nataly.

http://stemcell.ny.gov/faqs/what-dif...nd-multipotent

Quote Quote by caters View Post
with the advance in nanotechnology I would say sooner than 250 years for the tools. And with how fast we are advancing in cellular biology I would say sooner than 250 years for that.
What do you mean here? Bear in mind that sweeping speculative statements are against the rules.
caters
#8
Feb12-14, 10:03 AM
P: 38
Quote Quote by Ryan_m_b View Post
Is English your first language Caters? It's quite difficult to follow what you are talking about exactly.



Let me see if I've got this straight: you're proposing a system in which you have test tubes containing a reaction to replicate DNA. It's not just replicating the sequence but somehow building a functional chromosome with all the support proteins needed (histones for example). How do you propose this is possible? What is going on inside these tubes to achieve this? Furthermore you're proposing that each tube makes a different chromosome and you then are going to combine these into a, presumably empty, cell nucleus right?

Assuming I'm correct that this is what you mean what is the point if all this? At the end you're just going to have a cell which you could have just got by culturing the original one you harvested the DNA from. And where does totipotency come in here? Whether or not a cell is totipotent, pluripotent and what kind of cell it is isn't governed by the DNA sequence. Every cell has the same genotype, it's the complement of what genes are on and what are off that determine cell behaviour (simplistically, the microenvironment obviously plays a role).



So now you seem to be mixing in protein manufacturing but I don't understand what you mean here. In a separate system you want to PCR for a particular gene, then in a cell free manner produce large quantities of that protein. Whilst there are methods of doing this the most established is recombinant DNA technology. Regardless I don't see what this is meant to achieve. And what is a microtool?

So at the moment all you seem to have is a complicated way of replicating a cell without any real outlining of how you're going to do it (other than putting DNA in an empty nucleus, presumably with the cytosol and organelles intact) and an unrelated system building protein. What does any of this have to do with manufacturing totipotent cells?



You need to check your terminology. Pluripotent cells are defined as cells that can differentiate into any cell of the three germ layers (I.e. Any cell in an organism). A totipotent cell is the same but can also make placental cells which are not important post-nataly.

http://stemcell.ny.gov/faqs/what-dif...nd-multipotent



What do you mean here? Bear in mind that sweeping speculative statements are against the rules.
First off I did DNA extraction so I used proteinase K for the histones(I will form more histones when I get to the nucleus).

Now I will separate a pair of chromosomes after the DNA has replicated for a week.

Now the PCR I will do with the rest so that I have what is needed for forming proteins.

Centrifugation of the newly made protein from the rest follows that.

Than I put the supernatant in another test tube so that I can form a different protein.

It will fold right(the amino acid sequence says so)

I can do glycosylation.

The microtools will embed the protein into the membrane of the cell or organelle.
Ryan_m_b
#9
Feb12-14, 10:52 AM
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Sorry Caters but that's just a word salad. I suggest you go and look up processes like PCR to further your knowledge on what they are and what they can and can't do. Also proteins don't just fold because of their sequence, they need chaperones and specific environmental conditions. See gravenewworld's post for a further list of criticisms.

If you want to learn more about making stem cells I suggest properly looking into iPSCs and even adult stem cell research (pluripotent stem cells and even stem cells in general are not desirable for everything). If you want to learn about making cells from scratch I suggest looking into the nascent but promising field of synthetic biology. Above all though I'd work on improving your basic biology and communication skills.


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