BIO-360 BIOCHEMISTRY LABORATORY II
• XIII. WEEK
13th WEEK
ELECTROPHORETIC TECHNIQUES
PLASMID DNA ISOLATION
(APPLICATION)
Gel electrophoresis is a molecular
assay method widely used in the
detection of molecular weight,
amount and subtypes of purified
nucleic acid and proteins.
The advantage of the method is
that it allows the separation of
nucleic acid particles which
cannot be adequately separated
by other methods.
The most commonly used gel electrophoresis methods are agarose gel electrophoresis and
polyacrylamide gel
electrophoresis. For nucleic acids,
polyacrylamide gel
electrophoresis is generally used
for agarose gel and proteins.
http://www.creativebiomart.net/blog/principle-and-protocol-of-sodium-dodecyl-sulphate-polyacrylamide-gel-electrophoresis-sds-page/
PLASMIDE DNA ISOLATION
Molecular biology studies in the field of bacterial cells can be isolated from chromosomal DNA, plasmid DNA and phage DNA can be used as three different DNA.
Chromosomal DNA is always found in bacterial
cells, while plasmid DNA is an
extrachromosomal structure, it may not be
present in all bacterial cells.
PLASMIDE DNA ISOLATION
In bacteria, DNA is in the form of a double chain and a single ring. The structure called chromosomal DNA contains the genetic material of the bacteria. In addition, many bacteria have plasmid DNA carrying genetic information outside of chromosomal DNA. Plasmids can replicate themselves independently of chromosomal DNA and can be naturally generated by bacteria.
Plasmids are also found in Archaea and some
eukaryotic organisms. Plasmids can encode certain
genes and be transferred from one cell to another.
PLASMIDE DNA ISOLATION
Some plasmids are called sex factor and the
products of the genes carry the gene for the
bacterium to exchange genes with some other
bacteria and some genes are transferred. The
plasmids have a nucleic acid base sequence up
to about 1-200 kilobase pairs (kbp) in size and
are relatively small to chromosomal DNA.
There are three tandem stages in DNA isolation methods:
1) Cell homogenization.
2)Denaturation or proteolysis of DNA-protein complex and DNA degradation
3) Separation of DNA from proteins, RNA and other molecules by simple enzymatic and / or chemical methods.
In this experiment a plasmid with ampicillin resistance gene will be isolated!
https://en.wikipedia.org/wiki/Pla smid
