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PubMed1957

A chromatographic study of hydrolysis in the Feulgen nucleal reaction.

In a study on Feulgen hydrolysis of frozen-dried alcohol-fixed lily anthers, a chromatographic technique was developed to analyze the acid hydrolysate for some of the degradation products of nucleic acid. Hydrolysis was accomplished by 10 per cent perchloric acid at 20 degrees C., and a typical hydrolysis time-Feulgen intensity curve was obtained, with maximum staining occurring at 19 hours. Microphotometric measurements indicated that the amount of stain per nucleus was no different from amount in nuclei fixed and hydrolyzed by more conventional procedures. Uracil-containing material (from ribonucleic acid) was almost completely separated from thymine-containing material (deoxyribonucleic acid) of tissue sections by acid treatment for 1(1/2) hours. Adenine (purines), as the base, was effectively all removed from the deoxyribonucleic acid at the time of optimum hydrolysis. Detectable amounts of thymine-containing material appeared in the hydrolysate shortly after the onset of hydrolysis; and the amount increased rapidly with increased hydrolysis time. At the time of optimum hydrolysis approximately two-thirds of the total deoxyribonucleic acid thymine was lost. The removal of these thymine-containing fragments was linear with respect to time during the first 24 hours and occurred at a relatively high rate. Removal after 24 hours was also linear but was at a markedly lower rate. These results would suggest that two kinds of deoxyribonucleic acid exist in lily anthers; an acid-labile fraction amounting to approximately three-fourths of the total, and an acid-resistant fraction making up the remainder. In the Feulgen procedure much of the labile fraction is lost by the time of optimum hydrolysis and is not stained; most of the stable fraction remains in the tissue and is stained. In light of these findings the use of the Feulgen method as a means of determining cytochemically relative amounts of deoxyribonucleic acid in nuclei by measuring their Feulgen dye content was discussed.

Adenine↗
PubMed1958

The effect of HCl hydrolysis on the retention of thymidine in DNA.

Allium roots grown in C(14)-thymidine and H(3)-thymidine media were treated with N hydrochloric acid at 60 degrees C. as in standard Feulgen hydrolysis. The retention of the radioactive thymidine in DNA as a function of hydrolysis time was studied autoradiographically. No significant loss of label was detected until hydrolysis was extended beyond the optimal time for Feulgen staining. The data are consistent with the assumption that there is no significant loss of DNA during normal Feulgen hydrolysis in the material used.

Coloring Agents↗