Filter binding assay
Ø The term filter is used in to
describe a variety of different techniques in biochemistry, immunology,
virology and molecular biology.
Ø The molecular biology a filter
binding assay is used to characterized DNA protein interaction.
Ø This particular technique is also referred
to as filter assay or a protein binding assay and it can be used to identify
and characterised both DNA binding protein and the DNA sequence that can
interact with these proteins.
Ø Nitro cellular filter have been used
to decade to filter, sterilize solution.
Ø Filter binding assay discovered
accidently that DNA can bind to such nitro cellular filter because they lost
DNA preparation that`s why what is important is that nitro cellular filter can
indeed bind. DNA but only under certain condition. Single stranded DNA binds
radially to nitrocellulose filter, but double stranded DNA cannot bind.
Ø Protein does bind to nitrocellulose
membrane so it a protein is bounded to a double stranded DNA, then protein DNA
complex bind to nitrocellulose membrane.
Experiment – (Assay)
1.
Labelled
double stranded DNA poured through a nitrocellulose filter. The amount of
labelled DNA in the filter and filter bound material is measured which shows
that all the labelled doble stranded DNA material has passed through filter
into filtrate. This confirms that double stranded DNA does not bind to
nitrocellulose.
2.
A
solution of labelled protein is passed through nitrocellulose filter showing
that all the protein bound to the filter. This demonstrated that protein can
bind themselves to the filter.
3.
Now
double stranded DNA binds to protein and mixed with protein, to which DNA bind
to protein. Because of protein bind to filter the protein DNA-complex will also
bind and the radioactive is found bound to the filter rather than the filtrate.
4.
These
filters binding assay is direct measure of DNA protein interaction.
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