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.





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