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Dagl1 last won the day on January 7

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About Dagl1

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    Maastricht, Netherlands
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    Science; molecular biology (RNA and neuroscience (synaptic plasticity)), (quantum) physics, programming, behavioral psychology.
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    2-MSc biomedical sciences (molecular biology) Maastricht University --- Tohoku University
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    Molecular biology, cell biology
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  1. What qPCR kit (or just enzyme) do you use? It depends on both the primers in question (how easily do they form primer-dimers) and the enzyme used, however I generally used 250 nM (for both the forward and reverse primers). Optimal primer concentration can only really be obtained by titration, but I have only done this (once) when initial results were dubious/contained primer-dimer peaks (increasing annealing temperature wasn't really viable/did not help either). When titrating, Thermofisher (Sybr Green) recommends checking the 100-500 nM primer range, however other researchers sometimes check between 50 nM and 900 nM (as of researchgate). https://www.thermofisher.com/nl/en/home/references/protocols/nucleic-acid-amplification-and-expression-profiling/pcr-protocol/sybr-greener-qpcr-supermix-universal.html
  2. I suppose one way of thinking of it (which I am not able to defend with actual statistical data) is that smarter people may have a higher chance of finding an answer or new idea at any given moment, but that it is not a certainty. I am not sure if smart people will by definition come up with more ideas, but since that is a thought that pops up in your head, let's go with it: A smart person at any given moment has a higher chance than you to think of a good idea or some new knowledge, but that doesn't mean that said smart people will do so, therefore you can and probably should (if you want to create new knowledge) try, as there is always the chance that you will come up with a new idea that other people have not. The smarter people may outperform you in quantity, but there is a (almost) limitless amount of new knowledge that can be created, so just go for it, the chance that a smarter person will come up with THAT particular piece of new knowledge before you do is not hundred percent, so if you try enough, you may/will come up with new knowledge. Potentially many times someone smarter will have thought of it before you, so you try again, and again, each time rolling the dice and eventually you can win the jackpot (so to say). That said, I don't think being smart alone is a good determinant for creating knowledge either. Someone has to be interested in something, has to be thinking about the right stuff, at the right time, and combining already existing pieces of knowledge in just the right way to create a new piece of knowledge.T here seems to be some amount of 'randomness' to be involved in that process. People also have to be interested enough to write down and publicize there ideas for it to be a real addition to the common pool of knowledge. Additionally, a smart person may focus too much on a single subject to never come along a new piece of information, so by being interested and reading about many things, you increase the chance of coming up with something new anyway. You said you have a craving for knowledge, so if the above doesn't resonate with you, just go forget creating new knowledge for the sake of creating new knowledge. Instead, pursue finding/inventing new knowledge for the sake of bettering your own understanding of any given subject: in your pursuit of new knowledge, you will gain existing knowledge and new insights (that maybe other people have already had, but for you they will be new knowledge), and remember that every new piece of knowledge (in your mind) does bring you closer to the possible discovery of something truly new, unseen or unthought before. I found the book 'Where good ideas come from' by Steven Johnson, an interesting read about how new knowledge is (generally) created. I don't know how true the book is, but I feel it may be a perspective that can help you remove doubt from oneself (it is not about doubt in the slightest, but about the ideas that new knowledge generally only becomes available when several 'facts' or concepts known today, come together in someone's mind. That mind does not have to be a genius). I hope this helps, and if it does not, then I hope someone else can maybe provide you with something to reduce/eliminate your self doubt. PS. You can always go the Mohammed Ali route and just keep saying that YOU ARE THE GREATEST and that by definition YOU WILL CREATE NEW KNOWLEDGE (I do wonder about the efficacy of this method though).
  3. How would you verify the localisation after you get some data? Anyway I hope this helps you on your way with your essay, I am signing off for now.
  4. Sure! So we have a protein that is uncharacterized, meaning there probably are not going to be antibodies available. We can try producing proteins against it, but we could also try something else. What is cloning and what ability does it give us? How could we fuse these two things (fluorescence and cloning) together? The answer has a caveat though; there is of course a potential drawback to using fluorescence, in that it may not accurately describe localisation due to how we use fluorescence, but this is something that can be discussed/worked around later, and may not be in the scope of the question.
  5. What are ways that we can find out the localisation of a protein? How do we employ this by using cloning?
  6. I don 't know such reports, could you link some studies that show this? To your other question, a bit difficult to say since its pretty much unknown territory, I would imagine however that such transplants would make people quite sick due to unfamiliar bacteria, that normally don't live in the human gut micriobime, being added to that environment. I doubt people will suddenly become like tigers ^^.
  7. I don't know of any such studies. I also don't know if psychopathy actually increases chances to get offspring. Additionally there is no reason to think that psychopathy is something caused by specific genes, only that certain genes increase the risk (as of now, without enough evidence to support causal links, almost everyone has a chance to be(come) a psychopath, but certain gene variations can increase that chance). I don't see how 'brain drugs' are related to social engineering of non-psychopathic people (is it different for psychopathic people?). To your last question, no I haven't heard of such studies.
  8. I don't think evolution, especially for animals such as humans, where each generation takes quite a while, and which have extremely high inter-tribal (national) relations, will suddenly have more psychopaths in the genetic sense in the next hundred years. Is there any reason to think psychopathy is determined three or only a a few gene variations? About the fitness of a polygenetic trait that only increases fitness when all the right variants are in place: If they are not carried on the same chromosome, there inheritance is going to be a lot more difficult. I don't know about a way to simulate such a specific example. Also there are too many confounding variables to give a clear picture and to answer your other questions, at least for me.
  9. Per ml? or microliter? If you mean ml... then I think this is too low. In case you are talking about microliters, then 5nm/ul is still really low. You could attempt precipitation, but it also depends on the volume you have this amount in. I would always attempt using the same amount of RNA, so you may want to dilute the 80ng to 50ng in a reaction, and then using 50 ng of the 5ng/ul by taking a lot of volume (10 microl of RNA in your reaction). I am not entirely sure if that will be high enough for all subsequent procedures, but I think it could work. It is pretty low though, but I think it is worth trying, especially if you cannot get any new RNA from brain tissue. Maybe Charon can verify whether or not this really is the case.
  10. I am not sure about the relevance of the the HSA-miR palindrome, but my biochemistry knowledge is also not very up to date regarding how endonucleases cut exactly, maybe Charon can elaborate
  11. I think I may be misunderstanding, so apologies if I am missing the mark, but: in humans, each TSEN enzyme/subunit cuts one strand, and they don't seem to be directly complementary of each other. Also, a lot of mRNA is 'double-stranded' or forms hairpins/secondary structures, non coding RNA even more (I think). Important to note that mRNA normally binds to many mRNA-binding proteins, and therefore predicted structures (minimum free energy calculations) will often not actually form inside the cell (or be different from the predictions). The 5'UTR (Untranslated region) and 3'UTR of mRNA contains many structures as well. About 190 human genes (I think) contain an internal ribosomal entry site (IRES) within their 5'UTR, which can modulate ribosomal activity, these generally have the shape of hairpins as well. A viral enzyme of herpes virus, SOX, cuts at a single point, but recognises specific sequences in hairpins, so both structure and sequence is important. I don't really understand what you mean? https://journals.plos.org/plosbiology/article?id=10.1371/journal.pbio.1000307 https://journals.plos.org/ploscompbiol/article/figure?id=10.1371/journal.pcbi.1003517.g004 https://en.wikipedia.org/wiki/Insulin-like_growth_factor_II_IRES (couldn't find an article with eukaryotic IRES that shows it nicely)
  12. Imagine animals, such as apes, that live in tribes. Do we count a single tribe as a given population, or do we count all tribes as a given population. If we are talking about all tribes, then their size, frequency of inter-tribal breeding, and the amount of survival advantage provided by the trait, will determine the speed. Of course also the initial size of the given population is important. Simulations can sometimes account for these things, but it's quite a lot of work to tweak all such variables to mimic reality (I think). If you just want a broad idea, online simulators should be okay. I remember using toy models/simulations at university, but those were made in-house. You can play around with some simulation programs (important to run many simulations, as the outcome of each will be different): http://popgensimulator.pitt.edu/graphs/allele#! Or look for some other other gene frequency simulations. Maybe other people have more concrete answers (not sure if that simulator can do what you want).
  13. I may have to brush my knowledge about tRNA splicing, but could you elaborate what you mean by : I have been trying to find if SEN (TSEN in humans) tRNA splicing endonucleases actually cuts the opposite strand, but so far I have not found that they do: It definitely doesn't seem palindromic though. From quickly scanning these few articles it seems that for tRNA splicing there are two catalytic (TSEN2, TSEN34) subunits and two structural (TSEN15, TSEN 34). The last article (sci-hub link) shows this through mutating SEN (non-human version) 2 and 34 (see last picture). Please note that I have included the first article because it shows the structure of the complex quite nicely (I think), but that article is about specific mutations that cause pontocerebellar hypoplasia. Hope this is of interest/relevant to you! Articles (hopefully in order of the pictures): https://www.nature.com/articles/ng.204? http://www.genesilico.pl/rnapathwaysdb/Pathway/step/72/ https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4090602/ sci-hub.tw/10.1007/s00018-008-7393-y
  14. Dagl1

    Phage DNA

    I started explaining about viruses before I realised that phages specifically only talk about bacteriophages. I kept the information as you may find it useful or interesting anyway. The answer to your question, I would guess, is that it not great to do so for the phages. CRISPR systems may be one reason. Eukaryotic viruses (see below) can safely store their DNA within the nucleus, but this may be not feasible in bacteria. Instead even during latent periods they may only store their DNA within capsids. Sorry, I am not very familiar with phages;/ But based on this wikipedia article, I would guess that at least some viruses can stably integrate their DNA: https://en.wikipedia.org/wiki/Lysogenic_cycle This is in contrast to virus (please note I do not know enough about phages to say they do or don't have similar mechanisms) which don't immediately lyse their host cells. There are viruses which have a quiet phase, where they copy their RNA into DNA and store it close to the chromosomes within the nucleus. I am not very sure about the details/different types of latency, but some viruses can stay latent for super long periods of time. I think others can inhibit lysis for some time, thereby leading to larger amounts of viral particles. I am not sure if those that delay lysis (but do not become fully latent) also maintain viruses DNA as a stable molecule within the cell. I presume so, based on retroviruses existing, which insert copies into the chromosomal DNA of their host. https://en.wikipedia.org/wiki/Retrovirus Retroviruses such as HIV can also become fully latent, but recently I watched a video about Herpes virus (non retroviral) and how it escapes detection during latency:
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