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DeltaScience

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Everything posted by DeltaScience

  1. Well first one question? is it a protein or a gene? hehe because if you're runing a protein against a DNA marker it wont make any sense, althoug you could estimate the amount of a.a of your protein depending of the gene you're verifiyng but it´s harder. I think you can use a scale for the marker, if you say that it goes from 0 to 10 then you could use an scale in wich 10 would be a 100 or something like that. for the size of your protein you should check something about linear regresion using the MW as the reference for creating the equation, but I dont think that you have to use a plot to estimate the size of your protein, since I cant tell you the answer you should also check about the markers and what they're used for =P. I hope it was helpful. Good luck
  2. Well actually the citric acid is very important almost for every living organism, as they said here is an esential part of the krebs cycle, is also important for the regulation of the glycolisis, and I think in plants its most used as a bacteriocide, to protect the fruits and seeds from infections and as an atractive smell and flavour for birds and other animals to eat the fruit and so spread the seeds. Greets!
  3. Well I was using a special poylmerase to get a blunt end PCR product to use in the fusion PCR, the Takara PrimeStar wich is awesome, but then i run out of polymerase heheh and so far I have managed to get good amplification for my PCR product but only faint bands of my Fusion Product and then the transformation wasnt working, so I have the PCR product with blunt ends but now I need to fuse it with my vector wich is specially designed to fuse with the first primers I used for the PCR products, but this using the TaqPol hehe, thats my problem because i dont know if the extension time and final extension could be the problem with my reaction. Thanks
  4. Plasmids are just circular DNA wich can enter cells via different mechanisms like competence factors and etc, but the difference with viruses is that a virus can enter a cell or inject its DNA using specific membrane proteins, then the Viral DNA can be integrated into the cell genome, maybe you're confusing that some viruses can inject their DNA and it can stay as a plasmid inside the cell without integrating into the cell genome for some time, then it gets replicated and divided into new cells, and then after some time this Viral DNA can integrate the cells genome and start the infection, this depends on the cell culture conditions and the cell itself. Plasmids can also be integrated into the genome but its more likely that they will remain in their circular form and continue to be replicated with as the cells divides itself, this will continue until the cells doesnt require the genes encoded into the plasmid anymore. greets
  5. Hello everyone, well I'm performing some experiments using a new type of vector and for these I have to use Fusion PCR technique to get my genomic fragment into the vector, but I have been having troubles with these because my only Polymerase is the normal TaqPol haha, I think my problem lies in the Termocycler programing wich to be honest I dont know for this kind of PCR, I would really appreciate if someone could tell me the conditions needed for Fusion PCR with TaqPol, or some suggestions. Thank you. Greets
  6. Heyo everyone I'm delta science, I'm doing my bachelor in industrial and enviromental microbiology in Colombia, currently doing an internship in molecular biology and genetic engineering in germany, I also had some experience with bioprocess, production of bioethanol, biocontrolers for pest control, and some other stuff, I'm here to learn and also help if I can, see you on the other side people, have fun! +_+ Greets!
  7. Hi! well the explanation from aboyliketim is really good and accurate, I just wanted to share this video that I found in youtube wich I'm sure will give you a better perspective about this topic, just take into consideration of what aboyliketim told you and search for the structures!. I'm not the author of this video, no credits associated with me. Regards!
  8. it depends on what you need, normally this 2 parameters determine how reliably is your study, this is because the confidence interval will determine how much of your population will be represented in your model, since you have an unknown parameter and only a sample from the whole population, you need to guarantee that your unknown parameter will be inside this sample, thats why you need to set an confidence interval level to see how much are you taking, its called An interval estimate for a population parameter. Also the significance level will tell how big is your test, it represents a type of error, also called error type I, in wich it determines the posibilities for rejecting the null hypothesis when this one is true, so the smaller this error error the best, 0,05 is the standard, but ou can use 0,01, 0,5, it depends of your data. if your data is not too accurate or you see there's much error then you need to adjust this parameters in order to generate a valid model, even though the realiability of your model will be lower. I hope this is usefull. for further information you can check for this book, i think you can find it online: Applied Statistics and Probability for Engineers, the author is: Douglas C. Montgomery. Greets
  9. Hello!, well I dont know much about ion exchange cromatography but i do know that the isoelectric point of a protein determines it charge, and in this case it dependes basically on the pH of your elution buffer, for instance, for albumin (4,6-4,9) if you decrease the pH to a point lower than 4,6; then the protein will elute, and so on for the other proteins, also since in this case you're using KCL, you can try by using different pH or concentrations, the more the difference between the pi of a protein and the buffer you're using the more concentration of KCL you'll need to elute it. here have a look at this i found on the web, I hope its usefull for you: http://www.piercenet.com/files/TR0062-Ion-exchange-chrom.pdf I know it might not be the answer you're looking but hopefully will give you more usefull information. Regards
  10. Basically you need the plasmid to encode an essential gene for the microorganism, lets say for instance a plasmid encoding a gene to survive in a medium with an antibiotic, or a gene enconding a protein to degrade an essential substrate for the cell to grow, like lactose, and then keep the transformed microorganism under those conditions, this is like mandatory selection, you're forcing the cells to be under an specific condition in wich they need to use the genes that are present in the plasmid, this way you can guarantee that your cell wont loose the plasmid in the future, this is because it requires more energy for the cell to replicate this DNA. Also you need to take into account different parameters in order to get a succesfull transformed cell, like the size of your plasmid, if its too big it will be harder to get into the cells, and its also easier for the cell to loose it; what kind of genes it poses, if its circular or lineal, etc. I hope this helps to answer your question. greets
  11. Regarding the mitochondria evolution theory, it goes back to this endosymbiosis theory, in wich a single cell was fagocitated by a bigger cell, but instead of being degraded it became a sinergyc relation, in wich the fagocitated cell provided energy in exchange of protection and nutrients, also this is what it's said for chloroplasts as well, the difference lies in what kind of cells or microorganisms where fagocitated, it is said that mitochondria comes from the endocitosis of proteobacterias and the chloroplasts from the cyanobacterias, but its just a theory. that's a possible explanation about why the mitochondria and the chloroplasts has its own DNA and a double membrane. Regards
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