Welcome back to our second lecture on biotechnology.
In this lecture, we will apply some of the
techniques that we learned in the first lecture
and explore how DNA libraries are put together
as well as examine some DNA analysis techniques.
And finally we will look at some applications
of biotechnology in medicine and agriculture.
So by the end of this lecture, you should be
able to explain the benefit of DNA libraries as well
as compare RFLP and STR in DNA fingerprinting.
What are those you may ask? Well stay with
me and you will find out. By the end of this
lecture, you will also be able to discuss
some of the applications of biotechnology
in both agriculture and medicine.
Let us move into the beginning of genomic
libraries. Genomic libraries are similar to
our libraries except that there are volumes
of books that have overlapping sequences.
They are not independent volumes. There is
often some repeats in a DNA library. They
contain a representation of the entire genome
of an organism. The whole point is to store
that genome and fragments so that we can maybe
sequence it or figure out some of the genes
in the genome too. When we think about storing
these fragments in a library essentially all
we are doing is taking DNA fragments from
an organism's DNA workshopping them with restriction
enzymes. And then we will insert them into
a plasmid vector. We already know how that
works from the previous lecture and then we
take those vectors and ask bacteria to transform
and pick up these vectors and store them for
us in a nice neat library. Each piece of DNA
is in a different bacteria and there are perhaps
hundreds of bacteria to represent one genome
of maybe the fruit fly. Keep it mind, plasma DNA
is fairly small when we think about bacterial
plasmids. Artificial chromosomes have been
used to store larger fragments, but we will
move on to that later.
How do we get copies of eukaryotic genes?
This is really a good question because if
you recall from looking at our DNA transcription
and translation, how genes are expressed,
you might remember that some portions of the
DNA are expressed and some portions are not
in the form of introns and exons. The exons
are expressed, but bacteria don't have the
machinery to clip out the intron, the unexpressed pieces.
So we have to trick them into picking up just
the transcribable copies of DNA and this is
the way we do it by making a complimentary
DNA library. First of all, we will take the
eukaryotic DNA template and then we have transcription
occur. You know how that works by now. And
by that, we get a primary mRNA transcript
and then we need to clip out the introns in
order to just have the expressed sequence
where we see it in the mature messenger RNA.
Now that we have this mature messenger RNA,
we have the sequence of RNA that codes only
for the expressed portions of the proteins.
And in order to trick the bacteria to take
in all we need and just what we need, we need
to get a DNA copy of this. That is where the
complimentary DNA comes in. We will introduce
an enzyme called reverse transcriptase, which
guess what. Transcribes DNA from messenger
RNA, regular transcriptase goes forward.
Reverse transcriptase, we are taking a messenger
RNA molecule making a complimentary DNA strand.
Then we have single stranded complimentary
DNA and we need to get rid of the messenger
RNA template. So we will degrade those enzymes
and the messenger RNA will disappear and now
we need to introduce the DNA polymerase, the
enzyme that is going to polymerize the complimentary
strands. So DNA polymerase makes the complimentary
strand and now we have double-stranded complimentary
DNA with no introns ready to express the entire
protein. We can then introduce that into our