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Because T-RFLP relies on DNA extraction methods and PCR, the biases inherent to both will affect the results of the analysis. [ 6 ] [ 7 ] Also, the fact that only the terminal fragments are being read means that any two distinct sequences which share a terminal restriction site will result in one peak only on the electropherogram and will be ...
The DNA fragments produced by the digest are then separated by length through a process known as agarose gel electrophoresis and transferred to a membrane via the Southern blot procedure. Hybridization of the membrane to a labeled DNA probe then determines the length of the fragments which are complementary to the probe. A restriction fragment ...
Restriction digest is most commonly used as part of the process of the molecular cloning of DNA fragment into a vector (such as a cloning vector or an expression vector).The vector typically contains a multiple cloning site where many restriction site may be found, and a foreign piece of DNA may be inserted into the vector by first cutting the restriction sites in the vector as well the DNA ...
Short Tandem Repeat (STR) analysis is the modern day equivalent of RFLP. Not only does STR analysis use less of a sample to analyze DNA, but it also is a part of a larger process called Polymerase Chain Reaction (PCR). PCR is a process that can be used to quickly reproduce up to a billion copies of a singular segment of DNA. [3]
When removed from surrounding DNA by the PCR or RFLP methods, and their size determined by gel electrophoresis or Southern blotting, they produce a pattern of bands unique to each individual. When tested with a group of independent VNTR markers, the likelihood of two unrelated individuals' having the same allelic pattern is extremely low.
It is an extension to the restriction fragment length polymorphism (RFLP) method, using polymerase chain reaction (PCR) to more quickly analyse the results. Like RFLP, CAPS works on the principle that genetic differences between individuals can create or abolish restriction endonuclease restriction sites, and that these differences can be ...
This method yields high-quality, largely double-stranded DNA which can be used for both PCR and RFLP analysis. This procedure can be automated [9] and has a high throughput, although lower than the phenol-chloroform method. This is a one-step method i.e. the entire procedure is completed in one tube.