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Safety in Genetically Modified Organism - tRNA

  1. Oct 4, 2003 #1

    Another God

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    This idea just struck me. I think it might be one of those ideas which could be revolutionary, but at the same time, it could also just as equally be a stupid idea which everyone who knows what they are talking about can instantly see as a waste of time and money and completely impractical/unimplementable.

    Quick Background: DNA code is transcribed into mRNA, and those mRNA molecules are translated into Proteins according to the 'Universal Genetic Code' (which isn't completely universal) by tRNA's that are molecules which recognise codons in mRNA molecules. That is, according to a specific sequence in the tRNA (say AGC), it binds to the complementary RNA sequence (UCG) and adds the appropriate Amino Acid (Threonine) to the growing protein.

    Implication of this: If you change anyone of the tRNA recognition sequences, then the way DNA is translated changes completely. Any organism which you did this to would become completely inviable since the decription process of the DNA program has changed completely. It would be like taking a book and randomly substituting every letter, every space and every punctuation mark with a new one (even if we shifted every symbol one to the right say, and A = B, B = C etc), and then expecting the book to make sense. Chances are, every now and then a meaningful; word might pop up or something, but the sentence structure would disappear completely and so nothing meaningful could be done with it.

    The Idea:
    We could construct tRNA's so that all of the 64 possible codons are accounted for, accounted for in the same ratio, with a similar degree of efficiency of translation (this is important), but nonetheless slightly different. We could design this (probably through trial and error), and we could then get the genetic code for ..say....Wheat, and alter the code so that the mRNA molecules are the sequence required to produce the normal protein products, in light of our new tRNA's.

    In theory, the pant would produce the same proteins, the outcome would be the same, but the coding would be different.


    Why do this?
    Because one of the 'threats' associated with GMOs, is their escaping into the natural habitat. The chance of the actual plant itself escaping is effectively zero because our farm plants are selected to be great at providing food for us, not great at surviving (thats why 'weeds' kill our plants so easy...'weeds' are great at surviving). The threat comes from the fact that the pollen from the GMO's may accidentally cross pollinate into related wild plants, and those plants may in turn gain an advantage and out compete other wild plants.

    Another threat is the fear that the plant could cross pollinate into 'organic' plants of the same species in a nearby organic farm.

    By changing the tRNA's of GMO's, there is no chance of cross pollinating: The genetic codes are completely different. Any living organism could not use the DNA of these GMO's because it is meaningless to them. It uses a different language.

    Problems
    AS i said at the start, this idea only just occured to me. So now that I have written it out, a whole onslaught of problems have occured to me.

    changing the tRNA's is easy, but DNA does more than encode mRNA for codon translation....I am not sure how changing the DNA RE Codons might affect other signals imbedded in the DNA. Many promoter sequences and repressor binding sites are inside the DNA code of Genes....so by changing the code, we also change the expression rates inadvertantly.

    Also, doing this would actually mean re-engineeringn whole plants from scratch basically, and while this is sorta theoretically possible....Its not practical at all. I mean, its a few years worth of solid work from many many people no doubt. But then, the same thing was the case for the Human Genome Project... (Point in case: it toook how many years for them to 'sequence' a genome, let alone reconstruct one!!!!)

    Anyway, yeah, play with that. Monique and Ian, tell me if there are any more problems with it that I didn't consider.
     
  2. jcsd
  3. Oct 4, 2003 #2

    Another God

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    oopps, I got my universal code back to front. The mRNA sequence was UCG, so the encoded Amino Acid would have been Serine.
     
  4. Oct 4, 2003 #3

    Monique

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    Very interesting idea, AG, the problem is, as you stated, that altering the source code to fit the tRNAs would cause havok. Regulation of genes is very complicated and depends on non-coding sequences of the DNA which are very ill-defined.

    Computer programs are fairly good at finding exons and introns, but very bad at finding promotor regions. My former boss worked a lot with collagen, they had sequenced the complete gene which consists of 51 exons. At that time the sequence was not well known and computer programs had predicted the code, which was missing a lot of the exons!

    Just to illustrate that our predicting-knowledge is very limited. Predicting promotors regions is still extremely difficult.


    How would DNA polymerase be able to work? All the TATA boxes would be gone :)

    It is also thought that genes located in the same proximity somehow influence eachothers expression by conformational changes in the DNA.
     
  5. Oct 4, 2003 #4

    Monique

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    About the genetic code being universal, but not completely. I read that also not too long ago. Do you know any concrete examples?
     
  6. Oct 4, 2003 #5

    Monique

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    I was thinking: what if you just insert a GENE with the altered coding. That should work!!

    The only problem then is that the foreign tRNAs would interfere with normal translations, but that's what we've got protein engineers for! Genetically modify the RNA polymerase and the tRNA.

    But then there is a problem that other regions of the genome might code for genes with this new tRNA, just by chance..

    euhh

    engineer the polymerase such that ONLY the promotor of the inserted gene would be recognized :)


    Working with a gene at least makes things a lot easier than a genome, huh AG? :P
     
    Last edited: Oct 4, 2003
  7. Oct 4, 2003 #6

    Monique

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    You'll name me in your patent, right AG?
     
  8. Oct 4, 2003 #7

    iansmith

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    I don't see any problem but we could do better, add a 5th and 6th nucleotide. Scientist in Japan have actually add a 5th and 6th nucleodite. All we have to create is a organism with the rigth tRNA's or a code that requires 4 letter instead of 3. No organism on the planet can use this and Image of possible new protein you can create.

    What you have to think about is how good is your knowledge of the organism you are about to transform? All you have to do is transfrom every genes but you have to know in detail the DNA-binding sequence of repressor and promotors, know what the junk DNA does and you have to replace 30 to 50 thousand genes. That is one hell of a labor intesive project.
     
  9. Oct 4, 2003 #8

    Monique

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    I am sure they are working at it. At the department I used to work at, they were generating a protein-interaction map: a complete map of every signal protein interaction in a bacteria.

    Connecting all these interactions, one should be able to parse out metabolic pathways.

    Think about it, if they succeed, what information could be gained from it!


    Iansmith, I am aware of the extra nucleotides, good you mentioned it! It might work if the polymerase efficiently processes it.

    I see a problem with changing the length of codons, the whole genome transcription would be affected.

    Gaining information on all the junk-DNA seems like an impossible task.. 98% of the DNA is 'junk'! How would one go about approaching this problem?
     
  10. Oct 4, 2003 #9

    iansmith

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    Polymerase will not have a problem process it. During sequencing, polymerase add dd-NTP, and some mutagenic compound are similar to nucleic acid and are add during replication. It will probably not be a problem if a NTP can be add after the new base pair and so on.

    How would transcription be affected. There would only be more DNA to transcribed. Changing the length of the of genes by 33% will not matter but we have to make sure exon and introns are properly place and not tampered with.

    Maybe not study all the junk DNA but we have to keep in mind that changing the structure migth afftect the DNA-DNA and DNA-protein interaction which in term migth affterct the expression of certain proteins. Also, whole genome sequencing has some problem, they sequence genome in a certain set of condition. It is more obvious in bacteria. In our lab we have sequence genes and we have observe phase variation which cause protein to be truncated in iron replete condition but to be normal under iron restricted and in the addition of other signlas. The unfinish whole genome of our organism is available and we can see those truncated protein. Therefore some elements are design as having no sequence (Junk DNA) but it actually codes for protein under iron-restriction+misc. signals condition. What I am saying that we may id something as Junk but it migth be so usefull protein under a certain condition.
     
  11. Oct 4, 2003 #10

    Monique

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    Would it then be dangerous introducing such a base in the genome, or are you still planning on revamping the whole thing?

    So you ARE thinking about reinventing the whole genome..

    That is why I propose that it is not possible, at this point (maybe in 50 yrs) to set up an experiment that would deal with changing the sequence of the whole genome.

    Rather, it would be much more time and money effective to think about placing a single foreign gene in an otherwise normal genome.. that is how genetic engineering works anyway right?

    In this manner only all the regulatory proteins of this single gene are needed to be analyzed, the exact position of the promotor and first exon (which are difficult to define with algorithms).

    The only thing that needs to be figured out is how a novel RNA polymerase can be introduced that will only amplify that gene with the novel coding pattern.

    Otherwise all the interactions with the whole genome need to be analyzed, since we have 30.000 genes and every gene has an x number of proteins bound to it, how'd you figure out for all those interactions if they'd still work with the code changed? The task is too daunting..

    The human genome project, the sequencing is just reading of a code. The proteome-map is just the interaction of protein x with protein y. DNA-protein interaction is not that simple? You are going to footprint the 3 billion bases with the 30.000 proteins? And then test if they actually have a function? :D
     
  12. Oct 4, 2003 #11

    iansmith

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    I don't think the extra base would be dangerous.

    Why not? It's what AG was proposing, it would be a great challenge to be able to create an organism as simple as a bacteria. I am just playing with ideas, and try to excercise my mind. I think we had such a question/aasignment in one my advance genetic class.

    We trying to elimanate X-pollination. Not to duplicate what is being done. We need a novel approach. I know we won't solve the problem but it is fun to see what could be done.

    The problem is that the polymerase could be pass through X-pollination along with the gene.

    What I wonder is if we could create animal/plant plasmid/virus/transposon that would only propegate in a given species such as colE type of plasmid that only replicates in E. coli,
     
  13. Oct 4, 2003 #12

    Monique

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    Ah, you've got a point.

    So then tell me how you are going to determine all the regulatory units in the genome. If that is the way you plan on going :)
     
  14. Oct 4, 2003 #13

    iansmith

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    That it is one way to do it. I have to think about it in more detail. There is probably a DNA sequence that is only recognize by a giving species and that when X polination occurs it would not be pass along.
     
  15. Oct 4, 2003 #14

    Another God

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    But TATA boxes are prior to the translated sequence aren't they? They wouldn't need to be changed. The only thing that would need to be changed, under my idea, would be the translated region. Even the introns could be left the same ...in theory. Of course, you would need to figure out how to combine the right Genetic code for translation with the sequence which causes intron excision.... Actually, this wouldn't be a problem at all. =>

    What you do is ignore the INtron thing, write out the genetic code for the protein you want. You then search through that code for any sequence which matches that sequence recognised by the Intron excision process, and then stick an intron in after that with tha appropriate 'end of intron' sequence in it. Not hard really.

    Introns aren't a problem, the only problem remains with the gene internal promoters. (which even that mightn't be too big a problem since most of the main promoters are before the gene, and some are after....maybe if you change a gene which does require an internal promoter you can just supplement it by sticking a constitutive promoter infront of it, like a viral promoter...????
     
  16. Oct 4, 2003 #15

    iansmith

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    We got 30 to 40 thousand genes but it codes for way more proteins. Introns can be important and pose a problem if you do not take care of it. Depending on the promotor region, a gene can code for 2 different protein with different exon and introns.

    I also taught about another problem with reorganising the genome, post-transcritption modification. For example, we migth change the signal for cleaving or introduce of cleaving sequence.

    There also other problem with the strategy i propossed, post-translation modification and signal pepides.

    I went over my notes and mitochondrion and chloroplasst have their a different set of tRNA and do not go totaly by the universal code
     
  17. Oct 4, 2003 #16

    Another God

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    Yeah, I knew about the Mito and chloroplast genetic code, but even their code is close enough to the unviersal one that it seems quite apparent that the modern code is just the further refined version.

    One theory has it that originally there were only 2 base codons and it only coded for the most common amino acids, Proline, Alanine, Valine, Serine etc. Thats why you now see most of the Codons have a 'wobbly'(?) third base. The third base is always the least important.

    But yeah, what you say about Post translational modification, and then protein sorti....Hang on, nah, thats not a problem, because in the end the Protein should come out exactly the same. Any sorting signals in the protein will be exactly the same. Same as Post translational modification. unless the mRNA molecule interacts with the Protein molecule is has just encoded, nothing will change on the Protein side of things.
     
  18. Oct 5, 2003 #17

    iansmith

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    It does not matter if the protein but if you create new amino acid with the new base pair code then you migth have to change signal peptide and post-translational mod recognition in the enzyme.
     
  19. Oct 5, 2003 #18
    GMOs... all they ensure is less weeding. What's wrong with doing a bit of weeding?

    GMOz... its like creating a plastic bag by melting a plastic bag and making a plastic bag... redundant... whats the point?

    GMoZ... there are too many loopholes in genetically altering our food.

    Who's gonna know if a wheat gene has been transinfected by some looneytune megalomaniac with a Vrna that stimulates hyperdopemine or glutimates in certain sectors of the poplulation?

    Genetic engineering is too volatile a practice to put in the hands of humans, look what we did with DDT or 24D or PCBz etc... we're too stupid to handle the responsibility of mimicing lifes processes. In my opinion. Thank you.
     
  20. Oct 5, 2003 #19

    Monique

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    Quantumcarl, you are generalizing GMOs.

    Wouldn't you want to be able to grow crops in salt water, since fresh water supplies are not widely available and might be scarce in the future.

    Wouldn't you want to improve nutritional value, so that people in underdeveloped countries will have less protein-deficiencies?

    Wouldn't you want to improve efficiency of crops, so we'll need less space in the future, since the human population is expanding as crazy?

    Wouldn't you want to have crops that can grow on nutritionally-deprived grounds or toxic grounds, since who knows what the future might bring?

    Genetic engineering has been done for ages by humans, although low tech.. you don't think that maize is a natural plant do you? It was created by humans many centuries ago.

    Ofcourse I agree that we should be carefull with the genes we put from one organism in the other. But how unnatural it seems, it really is not.
     
  21. Oct 6, 2003 #20
    Compare the dangers of screwing up the genetic wheat pool (no pun here) to nuturing the fresh water resources we have and creating new ones. The reason most countries have lost their fresh water resources is because of mass deforestation and "global warming" or industrial emmissions.

    Wouldn't it make more sense to correct our own behavior than to genetically alter the behavior of some innocent bystanding organism like wheat or soy or flax?

    Maybe if the developed countries would leave the less developed countries to develope their resources rather than stripping their country side to develope better hamburger franchises, they would be able to take care of their own protein requirments without dangerously interfering with the complexity of genetic programing in plant organisms.

    Rather than place the emphasis of correcting our population problem on plants... perhaps it would be better to address our problem with stablizing influences like education and family planning.



    If we know what the future is bringing we can prepare for it by changing our behavior... not go around changing the behavior of everything else.

    Yes, it has survived through our selection of the type of plant best suited to our needs... not mechanically modified... selected over time. We let nature do the engineering.. which she is good at... we just selected what we liked best about her endevours. This left many different strains of maize still in the ecosystem... to fall back on in the event of a rust, virus or other disease... with genetically engineered plants, often the successors are completely erased by their aggressive features.
     
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