Sequence for Yourself Part I: Fragmentation of Chromosomes next
One Set of Chromosomes Displayed here are 23 pairs of chromosomes -- one copy of the human genome.
How is the genome sequenced? Let's do a little sequencing ourselves to find
out...
Current technology lets us read only 500 bases at a time, which means that we
can't start reading the A's, G's, C's, and T's at one end of a chromosome and
continue until we get to its other end. So the first thing we need to do is to
cut these chromosomes into small fragments.
Start with Many Sets of Chromosomes
To help us in piecing the fragments back together later on (explained in Part
V), the fragments need to overlap. To create the overlap, we'll have to cut up
many copies of the chromosomes, but at different locations.
We start with numerous cells from one person to obtain many sets of the
chromosomes.
Add Restriction Enzymes
Restriction enzymes will cut up, or cleave, DNA molecules wherever they
encounter a specific sequence of bases.
For example, one type of enzyme will cleave DNA wherever it encounters the
sequence AAGCTT.
Potential Cleavage Sites
Shown here are three identical chromosomes. The lines show all of the locations
that one type of restriction enzyme might target.
However, we don't want the enzymes to cut at every possible location because
then there would be no overlap.
Limiting Cleavage
To limit the cutting, we need to use a low concentration of the enzyme. With a
low concentration, there just isn't enough of the enzyme available to cut the
DNA at every possible site.
What we end up with are many overlapping fragments. At this point each fragment
is about 150,000 base pairs long.
One Set of Chromosomes
Start with Many Sets of Chromosomes
Add Restriction Enzymes
Potential Cleavage Sites
Limiting Cleavage