Chargaff’s Experiment and Why is Chargaff's Rule Important?

Let’s begin with Chargaff’s rule.

Chargaff’s rule states that

Rule 1: The amount of Adenine ~equals the amount of Thymine

The amount of Guanine ~equals the amount of Cytosine

The amount of purine = the amount of pyramidine

Chargaffs Rule Summary

Rule 2: The amount of A+T ≠ amount of G+C. This ratio varies among different organisms but same in different tissues of the same organism 

Chargaff’s Experiment

He developed a paper chromatography technique to separate and quantify nucleotide bases

Chargaff's experiment

Let us divide his experiment into two parts

Experiment. 1: Determination of base composition of DNA from different species

First he isolated DNA from different organisms like Human tissues, E.coli, rat etc. separated and quantified nucleotide bases

Experiment 2: Determination of base composition from different sources of same organism

First he isolated DNA from different tissues of the same organisms like DNA from Human thumus, spleen, sperm etc and quantified nucleotide bases


His procedure consisted of three steps.

  • The first was the separation of the DNA mixture into individual components by paper chromatography.
  • Next,the separated compounds were converted into mercury salts.
  • And finally, the purines and pyrimidines were identified via their ultraviolet absorption spectra.

From the result he derived Chargaff's Rule

Watch this video for better understanding

Why is Chargaff's Rule Important?

Explained base-pairing regularities or "complementarity relationships" of DNA thus providing the most important clue for the Watson and Crick for deducing the structure of DNA. At the beginning, Watson and Crick proposed “Like with like” base pairing where purine pairs with purine and pyramidines with pyramidines; A=A, G=G etc.  Complementary base pairing provided the most stable orientation of base pairs in the DNA double helix. This orientation (purine to pyramidine base pairing) best matched with the X-ray crystallographic data. Thus Chargaff’s Rule provided the solution for solving this puzzle.

DNA copying mechanism

Watson and Crick could explain how DNA makes copies of itself using this base complementarity relationship. One DNA strand serves as a template for the synthesis of other strand.


  • Tetra nucleotide Hypothesis by Phoebus Levene
  • Triple helical structure of DNA  proposed by Linus Pauling
Learn More: How to calculate the percentage of bases  in a DNA strand using Chargaff’s rule? (Chargaff's Rule Questions)

                       Watson and Crick 1953 paper

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