How to isolate DNA from human blood at home?

By Anonymous -

 






DNA extraction is an essential technique in molecular biology. Isolation and purification are important to determine the unique characteristics of DNA , including its size, shape and function. Isolation of high-molecular weight DNA has become very important  with the increasing demand for DNA fingerprinting, restriction fragment length polymorphism , construction of sequencing libraries and PCR analysis in research laboratories and industry.

Principle:

DNA extraction of DNA involves three main steps cell lysis ,protein separation and DNA purification.Cell lysis is usually performed by incubation of cells in buffer  having protease and detergent.





Equipment required:

Centrifugation for 15 ml faclon tubes
Microcentrifuge for 1.5 ml tubes
Adjustable micropipettes 1 ml and 200ml

Reagents required:

  •  TEN buffer
  •  TE buffer
  •  10 percent SDS
  •  6M NaCl
  •  Protinease -K solution 20 mg
  •  Phenol -Choloroform -Isoamyylalcohol
  •  Absolute Ethanol
  •  75 percent Ethanol
  •  Low TE buffer

Precaution to be taken during DNA extraction:

  • Make sure that vials and tips are DNase free.
  • Avoid vortexing during DNA isolation.
  • You have to make sure that the extracted DNA is treated with RNase to remove RNA.It is good to use columns to clean the DNA .
  • Ship the DNA samples on Dry ice.


Procedure:


1. First add 1ml TE buffer into 200ul blood and

 mix it very well.

2. Centrifuge at 4000rpm

3. Discard the supernatant and add900 ul TE

 buffer.

4. Centrifuge at 4000rpm.

5. Discard the supernatant and add 800 ul TE

 buffer.

6. Centrifuge at 4000 rpm.

7. Discard the supernatant and add 200ul TEN

 buffer, 20ul SDS and 10ul proteinase-k.

8. Incubate at 56 degree.

9. Place the tubes on ice and add 6M nacl.

10. Centrifuge at 4000 rpm for 15 min.

11. Transfer the supernatant in a freshly labeled

 tubes.

12. Add equal amount of chilled isopropanol.

13.

Centrifuge at 8000 rpm.

14. Add 200ul absolute ethanol

15. Centrifuge at 8000 rpm.

16. Add 200ul ethanol.

17. Centrifuge at 8000 rpm.

18. Discard the supernatant and add 100ul Low

 TE buffer.

19. Store DNA.


Result:


Presence of DNA can be confirmed by

electrophoresing on an agarose gel containing

ethidium bromide.





Comments

Post a Comment

Popular posts from this blog

How did the viruses originated?

By Anonymous -
Image
  An infective agent that typically consist of a nucleic acid molecule in a protein coat, is too small to be seen by light microscopy, and is able to multiply only within the living cells of a host. u   Latin word u   Meaning poison u   Smaller than bacteria u   Smaller than the smallest cell Can you find the size of viruses? u   The name virus was given by Beijernick in 1898 u   He studied filtered  plant juices and    found that  they caused  healthy plants to    become sick . 1.       Viruses have non- living structures 2.       Non cellular structure 3.       Contain a protein coat called the capsid 4.       Viruses have nucleic acid core containing DNA or RNA 5.       Capable of reproducing only when inside a host cell 6.       Most viruses infect only specific host cells 7.       Viruses are inactive outside of the host cells 8.       Viruses cause many common illnesses/diseases 9.       Some viruses may cause some cancers like leukemia Edward Jenner introduced the term virus in
Read more

How the spoilage food effects the human health?

By Anonymous -
Image
      Spoilage food more commonly referred to as food poisoning, is the result of eating contaminated, spoilage or toxic food. The most common symptoms of food poising include nausea, vomiting and diarrhea. Although it’s quite uncomfortable, food poisoning is not unusual. Food spoilage and deterioration is no accident. It is naturally occurring process. To understand how to maintain the quality of food and prevent spoilage, we need to know what can cause it. Factor that effect food spoilage. Microorganism Enzyme Air Light Insects Temperature Time     Preservative frequently involves preservation the growth of bacteria, fungi and other microorganisms. A number of methods of preservation can be used that can either totally prevent, delay or otherwise reduce food spoilage. Detection of spoilage food: The detection of food spoilage may include an appearance different from fresh food, change in color, a change
Read more

Why heart attack is increasing among the young people?

By Anonymous -
Image
  It was rare for anyone younger than 40 to have a heart attack .Now 1 in 5 heart attack patients are younger than 40 years of age.Having a heart attack in your 20s or early . The rise in heart attack among younger people can largely be attributed to lifestyle .Overbooked schudele, pressure to compete ,constant stress and anxiety, overeating and self –medicating with alcohol and drugs take a toll on the heart. Hypertension or high blood pressure is one of the biggest trend factors for cardiovascular disease .inappropriately just like the trend in heart attacks, the occurrence of hypertension is rising faster in young adults than in older adults. High blood pressure makes your heart muscles thicker, harm your blood vessels and increases your risk of a heart attack.     Without warming,your heart stops due to an electrical disturbance that causes an abnormal heart rhythm.Heaert attacks increase the risks of sudden cardiac arrest,which can cause death without immediate treatment.
Read more