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What are the differences between conventional pcr andreal time pcr?
PCR allows amplification of DNA for a specific gene, after too many cycles of PCR the result will reach saturation, basically meaning all of the DNA has been amplified. Conventional PCR will basically tell you whether or not a gene is expressed in your sample. This can be done semi-quantitavely if the PCR is performed for a low number of cycles, ie it will tell you whether one sample expresses more of your gene of interest than another sample. The results are seen by separating the PCR products by agarose gel/ethidium bromide electrophoresis. Real-time PCR will record exactly what cycle of PCR a detectable level of amplified product became detectable, giving a far more accurately quantifiable estimation of gene expression.
What is the difference between Clean-room and sterile-room?
You may want to double-check this, but I believe PCR-clean simply means that there are no DNases or RNases on the item, but they still could have nucleic acid on them. Essentially there is nothing on them that would interfere with nucleic acid amplification achieved with PCR, but any genetic material they may have will be amplified. Sterile means that there is absolutely no genetic material on the item itself (usually achieved via autoclaving where the temperatures climb so high that they would denature the DNases and RNases anyway). Nutshell: PCR-clean = wiping with RNA away and DNA away Sterile = bleaching and autoclaving
Why 10x buffer use in pcr?
It provides a suitable chemical environment for optimum activity and stability of the DNA polymerase.
What are the steps in a direct-sequencing experiment?
Lyse cells, purify DNA, amplify genes by PCR, and insert genes into plasmid
Why are scientists so interested in extremophiles?
Extreme environments have been useful to scientists in inventing PCR. It was in an extreme environment like the geysers of Yellowstone that a scientist discovered that a bacteria was living in the extremely hot water and yet still could function. Before PCR we knew we could separate a strand of DNA by heating it, but there was no polymerase to duplicate it that would work at such a high temperature. The bacteria in the hot water had a polymerase that would. So now scientists use that to do PCR and create many copies of DNA.
