, Volume 7, Issue 2, pp 87-94

A simple and efficient procedure for isolating plant chromatin which is suitable for studies of DNase I-sensitive domains and hypersensitive sites

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Summary

A simple and rapid procedure has been developed for the isolation of chromatin from plant leaves. The molecular weight of the DNA extracted from these chromatin preparations is comparable to that of DNA isolated by a conventional purification procedure (CTAB-CsCl-method). These results suggest that almost no degradation occurs during the isolation procedure. The effect of DNase I on three different groups of genes was studied; one of them, encoding the NADPH-protochlorophyllide oxidoreductase (PCR), represents a gene which is actively transcribed in etiolated leaf tissue. The other genes examined encode the hordein seed storage protein and 26S ribosomal RNA. The hordein genes are known to be inactive in leaves.

The hordein and rDNA genes were found to be resistant to low levels of DNase I, while the gene for the PCR was highly sensitive to DNase I. During the course of digestion of the PCR gene, discrete cleavage products are generated. These indicate the presence of DNase I hypersensitive sites in the vicinity of the PCR gene in etiolated leaves. As a control ‘naked’ DNA has been digested with DNase I. No differences in sensitivity between the PCR gene and the hordein genes can be detected.