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Nucleic Acids Research, 1994, Vol. 22, No. 25 5702-5708
© 1994

Articles

Detection of circular excised DNA deletion elements in Tetrahymena thermophila during development

Meng-Chao Yao* and Ching-Ho Yao

Division of Basic Sciences, Fred Hutchinson Cancer Research Center 1124 Columbia Street, Seattle, WA 98104, USA

*To whom correspondence should be addressed

Received August 9, 1994. Revised November 4, 1994. Accepted November 4, 1994.

Extensive programmed DNA deletion occurs in dilates during development. In this study we examine the excised forms of two previously characterized deletion elements, the R- and M-element, in Tetrahymena. Using divergently oriented primers in polymerase chain reactions we have detected the junctions formed by joining the two ends of these elements, providing evidence for the presence of circular excised forms. These circlar forms were detected in developing macronuclear DNA from 12 - 24 h after mating began, but not in mlcronuclear or whole cell DNA of vegetative cells. They are present at very low abundance, detectable after PCR only through hybridization with specific probes. Sequence analysis shows that the circle junctions occur at or very near the known ends of the elements. There Is sequence micro-heterogeneity in these junctions, which does not support a simple reciprocal exchange model for DNA deletion. A model Involving staggered cuts and variable mismatch repair is proposed to explain these results. This model also explains the sequence micro-heterogeneity previously detected among the junction sequences retained in the macronuclear chromosome.


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