QUALITATIVE AND QUANTITATIVE ANALYSIS OF DNA BY SPECTROPHOTOMETRY

 

ABOUT AUTHORS:
Amar Nagesh Kumar1, Reena jose2, Praswetha Reddy3
1 Lecturer, Department of Biochemistry, SSR Medical College, Mauritius
2 M. Sc Biotechnology, Yogi Vemana University
3 M. Sc Biochemistry, Mahatma Gandhi P.G College, Guntur

OBJECTIVE:To estimate the given samples of DNA qualitatively and quantitatively and know its purity and concentration.

 BACKGROUND
Quantification of nucleic acids is commonly used in molecular Biology to determine the concentration of DNA and RNA present in the mixture As subsequent reaction or protocols using the nucleic acids sample of a required particular amount of optical performance. Both DNA and RNA exhibit strong absorbance of UV due to the presence of conjugated double bonds of the constant purine and pyramidine bases and these have characteristics of OD (optical density)of absorbance maximum at 260nm which is linearly related with the concentration  of the DNA in the solution up to the OD value of 2 . The spectroscopic method is used to check the purity of DNA. Proteins are the major contaminants in the nucleic acid extracts and these have the maximum absorbance at 280nm. Value less than 1.8 signifies the presence of proteins as impurities.

REFERENCE ID: PHARMATUTOR-ART-1134

MATERIALS & METHODS
Materials required
· SSC buffer(saline sodium citrate)pH 7
· Quarts cuvette
· Distilled water
· Tissue paper
· Conical flask
· Beaker
 UV spectrophotometer

REAGENT  PREPARATION:  
SSC BUFFER  20ml
* 0.3Mtrisodiumcitrate 1.7 gm dissolved in 20ml of distilled  water is 0.3 tri sodium citrate
* 3M sodium chloride
* 50 gm dissolved in 20ml of distil water is 3M sodium chloride

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Comments

how the DNA is diluted 100 times?

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