Heterogeneity of ribosomal gene methylation in inbred lines

We have surveyed the ribosomal DNA (rDNA) of a number of inbred lines of maize with the methylation-sensitive restriction endonuclease Hpall. Although a large portion of the rDNA is not accessible to HpaII cleavage, MspI (which has the same recognition sequence as HpaII, but a different methylation' specificity) produces an even distribution of rDNA fragments, indicating that the uncut fraction is due to methylation and not lack of the recognition site. A fraction of maize rDNA is always accessible to Hpall digestion. The characteristics of the Hpall-sensitive fraction varied among inbred lines of maize (see Fig. 1 and Table 1). HpaII digestion of maize rDNA produced either one repeat unit (RU) band, two bands, or three bands. In cases where two or three bands were observed, one fragment was of RU length and additional fragments were lower in molecular weight by 1.1 kbp and 2.3 kbp. B37N was the only inbred line in which all three bands were observed. All other inbred lines exhibited either a one- or two-band pattern. In lines with two bands, only the RU length band and the smallest band were present. Lines with a single band had only the RU length band present. The results from this survey of inbred lines show that not all rDNA repeat units in a maize genome are structurally identical with respect to methylation at Hpall sites. In addition, the number of hypomethylated Hpall sites present in the rDNA RU varies among inbred lines of maize. Experiments are being conducted to determine the functional significance of the observed structural heterogeneity in ribosomal gene methylation.

Table 1. Inbred lines of maize surveyed with HpaII
 
Line Number of Bands Present Size of Bands(kbp)
B37N 3 9.1, 8.0, 6.9
BMS 2 9.1,6.9
K10 2 9.1,6.9
Ohio Pop 2 9.1,6.9
Tx303 2 9.1.6.9
WKF 2 9.1,6.9
B73 1 9.1
GF 1 9.1
Mo17 1 9.1
Ny302 1 9.1
Tx601 1 9.1

Figure 1. Maize DNA isolated from leaves (Zimmer et al. Plant Mol. Biol. Newsl. 2:93, 1981) was digested with either HpaII or MspI, and fragments were separated by electrophoresis on 0.8% agarose gels. Fragments were transferred to nitrocellulose by Southern blotting and probed with a soybean rDNA probe containing the entire repeat unit (pGmr1). The three bands produced by HpaII digestion of B37N were 9.1, 8.0, and 6.9 kbp in size, respectively.

Eldon R. Jupe and Elizabeth A. Zimmer
 
 


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