Structure and variation in ribosomal RNA genes of pea

Abstract

A complete ribosomal DNA (rDNA) repeat unit has been cloned from the genome of Pisum sativum (garden pea) and used to construct a map containing a total of 58 cleavage sites for 23 different restriction enzymes. Regions encoding 18s and 25s ribosomal RNA (rRNA) were identified by R-loop analysis. A 180 bp sequence element is repeated eight times in the intergenic 'nontranscribed spacer' (NTS) region, as defined by eight evenly spaced RsaI cleavage sites. Sequence heterogeneity among these elements (subrepeats) is indicated by the presence of an NcoI site within the five RsaI subrepeats distal to the 25s rRNA gene but not in the three subrepeats proximal to this gene, and also by the presence of an additional RsaI cleavage site in one subrepeat.The approximately 4000 copies of the rDNA repeat in the pea nuclear genome show considerable heterogeneity with respect to the length of the NTS region, and differences are also frequently observed between different genotypes. In both cases the length variation appears to be due primarily to differences in the number of subrepeat elements.Comparison of rDNA restriction maps for two pea genotypes separated for hundreds or perhaps thousands of generations reveals that they contain many rDNA identical repeat units. This data is consistent with the view that new rDNA variants are fixed only infrequently in the evolution of a species.Differences also exist between the rDNA repeats of a single genotype with respect to the degree of base modification at certain restriction sites. A large number of sites known to exist in the pea rDNA clone are not cleaved at all in genomic rDNA, or are cleaved in only some copies of the rDNA repeat. We believe these examples of incomplete cleavage results mostly from methylation, although it is difficult to rule out the possibility of sequence variation in all cases. Most putative modifications are best interpreted in terms of cytosine methylation in CG and CXG sequences, but at least one example is more consistent with adenine methylation.We also have constructed a more detailed restriction map of the wheat rDNA clone pTA71 and present a comparison of this map to our map of pea, pumpkin, and wheat in order to assess the amount of useful evolutionary information that can be obtained by comparison of such maps.

Single-stranded replication intermediates of ribosomal DNA replicons of pea.

“…The larger repeat, located on chromosome 7, is more abundant representing -3000 tandem copies or a total of 27 000 kb (Polans et al, 1986;Jorgensen et al, 1987). There are -900 tandem copies or -8000 kb of the smaller repeats on chromosome 4 (Polans et al, 1986;Jorgensen et al, 1987). The repeats are located at or near the nucleolar organizers on their respective chromosomes (J.…”

“…The repeats differ in size because of the number of 180 base subrepeat units located in the spacer region. The 9 kb repeat also has two EcoRl sites in the spacer region which are absent in the 8.6 kb repeats (Jorgensen et al, 1987;Van't Hof et al, 1987). The EcoRI fragments from the 9 kb repeats are 0.190, 1.3, 3.9 and 3.7 kb, those of the 8.6 kb repeats are 3.7 and 4.9 kb.…”

Extensive Conservation of Linkage Relationships Between Pea and Lentil Genetic Maps

“…Pea DNA probes included the 45S ribosomal DNA repeat (pHA2, Jorgensen et al 1987), genes coding the plastid-specific, cytosolic, and nodule-specific glutamine synthase (pGS185, pGS299, pGS340, Tingey et al 1987), the plastid-specific glyceraldehyde 3-phosphate dehydrogenase gene (pNcol, Cerff et al 1986), and the gene coding the chloroplast ribosomal protein-22 (rpl22, Gantt et al, 1991). A plasmid containing an alfalfa leghemoglobin sequence (pALb, Dunn et al 1988) and one containing a soybean actin gene (pSAc3, Shah et al 1982) also were used in the study.…”

Genotypic variation within asexual lineages of Taraxacum officinale.

“…The rDNA restriction site changes in the Taraxacum offspring are located about 1300 bp from the EcoRI site at the 3' end ofthe 25S gene. This region of the ribosomal cistron is known to consist of subrepeating elements that contribute to rDNA repeat length variation in several plant species (30)(31)(32)(33)(34)(35). In Taraxacum, length polymorphisms ranging from 100 to 1000 bp map to this region (8).…”