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Name: Karen L.
Status: Student
Age: 18
Location: N/A
Country: N/A
Date: March 2004


Question:
How is reverse transcriptase used in molecular biology? and could you give me an example?



Replies:
Reverse transcriptase copies RNA to DNA. It is often used to make a "library" of DNA clones from RNA. Let's say you are interested in studying only the genes that are expressed in a newly-fertilized frog embryo. First, you isolate the messenger RNA from the embryo. Now you use reverse transcriptase to make DNA copies of the RNA. Then you clone the DNA pieces into some type of vector, such as a plasmid or phage. Now you have a stage-specific library. Every clone in the library is there because it encodes an RNA that is expressed in your newly-fertilized embryos. This is very different from a total genomic library, which should represent all of the DNA in the nucleus, regardless of when it is expressed.

Paul Mahoney, PhD


There are lots of uses, but one simple example is reverse transcribing an mRNA message back into DNA. mRNA is very unstable, but it has already been processed, ie. it has had the introns removed. So it is a complete protein "recipe" without the interrupting sequences in it. Reverse transcriptase is used to turn it back into DNA but without the introns. This DNA is called cDNA or copy DNA. Then it can be used for study. For example one thing they use copy DNA for is as a "probe" for finding where in the genome that mRNA came from. If the DNA is made to be single stranded, the half that is opposite will stick to the place in the DNA that the gene is found.

vanhoeck



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