DNA sequencing using gel electrophoresis

M

mariyam95

Messages
114
Reaction score
36
Points
38
Hi :)
i need help in this topic plz if u can post any notes and links that might help me coz im seriously desperate now :(:cry:
thanks in advance!
 
Haider2011

Haider2011

Messages
58
Reaction score
7
Points
18
this may help u in all application topics
 

Attachments

  • 1. Protein Chemistry.pdf
    329.1 KB · Views: 25
  • 1. Stucture Determination.pdf
    228.5 KB · Views: 18
  • 2. Genetic Information.pdf
    293.5 KB · Views: 18
  • 2. Separation Techniques.pdf
    311.4 KB · Views: 18
  • 3. Energy.pdf
    95.2 KB · Views: 19
  • 4. Metals in biological systems.pdf
    81.4 KB · Views: 19
KurayamiKimmi

KurayamiKimmi

Messages
191
Reaction score
80
Points
38
mariyam95 said:
bio :(
Click to expand...
okies here it is:)

Electrophoresis:
It is a technique that is used to separate DNA fragments of different sizes .
use:
It is used for genetic profiling and for investigating the sequence of bases in a particular length of DNA

Preparing the DNA sample:
  • A region of DNA that varies from person to person is chosen , this is known as variable number tandem repeats -VNTR. Only identical twins have the completely same VNTRs.
  • The DNA is extracted from the person/crime scene,etc and then the quantity of DNA is increased by the 'polymerase chain reaction' which makes many copies of the DNA therefore even if you start with only a single cell you can have enough sample for electrophoresis.
  • The DNA is chopped into pieces using restriction enzyme that are known to cut the DNA close to VNTR sections.
Process of electrophoresis:
  • A tank is filled with agarose gel which is very pure, and a direct current is passed continuously through the gel.
  • The DNA fragments are all negatively charged due to how they were cut, and so are attracted towards the anode (+ve electrode) and so travel to it , the smaller ones are faster .
  • When the current is switched off, the gel contains DNA fragments that have ended up in different places , but this is not visible straight away.
Analyzing the fragments:
  • To make them visible, they are first transferred very carefully onto absorbent paper , which is placed on the gel.
  • The paper is then heated just enough to make the DNA's two strands separate from each other. Short sequences of single stranded DNA called probes are added, which contain a radioactive isotope. These pair up with the separated DNA strands on the paper , as they have complementary base sequences to VNTR regions.
The paper if now placed on an X Ray film , the radiation emitted by the probes makes the film go dark. So we end up with a pattern of dark stripes on the film, matching the positions that the DNA fragments reached on the gel.

(Do keep in mind that although I've written it in point form , don't do so in the exam...)
There's also a process called 'Southern blotting' do let me know if you'd want that.:)
 
M

mariyam95

Messages
114
Reaction score
36
Points
38
KurayamiKimmi said:
okies here it is:)

Electrophoresis:
It is a technique that is used to separate DNA fragments of different sizes .
use:
It is used for genetic profiling and for investigating the sequence of bases in a particular length of DNA

Preparing the DNA sample:
  • A region of DNA that varies from person to person is chosen , this is known as variable number tandem repeats -VNTR. Only identical twins have the completely same VNTRs.
  • The DNA is extracted from the person/crime scene,etc and then the quantity of DNA is increased by the 'polymerase chain reaction' which makes many copies of the DNA therefore even if you start with only a single cell you can have enough sample for electrophoresis.
  • The DNA is chopped into pieces using restriction enzyme that are known to cut the DNA close to VNTR sections.
Process of electrophoresis:

  • A tank is filled with agarose gel which is very pure, and a direct current is passed continuously through the gel.
  • The DNA fragments are all negatively charged due to how they were cut, and so are attracted towards the anode (+ve electrode) and so travel to it , the smaller ones are faster .
  • When the current is switched off, the gel contains DNA fragments that have ended up in different places , but this is not visible straight away.
Analyzing the fragments:

  • To make them visible, they are first transferred very carefully onto absorbent paper , which is placed on the gel.
  • The paper is then heated just enough to make the DNA's two strands separate from each other. Short sequences of single stranded DNA called probes are added, which contain a radioactive isotope. These pair up with the separated DNA strands on the paper , as they have complementary base sequences to VNTR regions.
The paper if now placed on an X Ray film , the radiation emitted by the probes makes the film go dark. So we end up with a pattern of dark stripes on the film, matching the positions that the DNA fragments reached on the gel.


(Do keep in mind that although I've written it in point form , don't do so in the exam...)
There's also a process called 'Southern blotting' do let me know if you'd want that.:)
Click to expand...
thank you ...but plz can u help me with DNA sequencing ?
 
You must log in or register to reply here.
Top