Abstract Double-strand break (DSB)-induced gene con- version in yeast was studied in crosses between ura3 het- eroalleles carrying phenotypically silent markers at ap- proximately 100-bp intervals, which allow high-resolution analyses of tract structures. DSBs were introduced in vivo by HO nuclease at sites within shared homology and were repaired using information donated by unbroken alleles. Previous studies with these types of crosses showed that most tracts of Ura + products are continuous, unidirectional, and extend away from frameshift mutations in donor al- leles. Here we demonstrate that biased tract directionality is a consequence of selection pressure against Ura – prod- ucts that results when frameshift mutations in donor alleles are transferred to recipient alleles. We also performed crosses in which frameshift mutations in recipient and do- nor alleles were arranged such that events initiated at DSBs could not convert broken alleles to Ura + via a single gap repair event or a single long-tract mismatch repair event in heteroduplex DNA. This constraint led to low recombina- tion frequencies relative to unconstrained crosses, and in- hibited preferential conversion of broken alleles. Physical analysis of 51 DSB-induced products arising from multi- ple recombinational repair events suggested that hDNA formation is generally limiting, but that some hDNA re- gions may extend more than 600 bp. Among these prod- ucts, markers separated by 20 bp were independently re- paired about 40% of the time. Key words Gene conversion · Double-strand breaks · Heteroduplex DNA · Yeast Introduction Gene conversion is the non-reciprocal transfer of informa- tion from a DNA duplex to a homologous duplex, a pro- cess that has been widely studied in yeast (reviewed by Petes et al. 1991). Conversions have been explained by two types of models. One type proposes heteroduplex DNA (hDNA) intermediates with conversion resulting from the correction of mismatched bases in hDNA (Holliday 1964; Meselson and Radding 1975; Radding 1982). Double- strand break (DSB), or gap repair models (Szostak et al. 1983; Thaler and Stahl 1988; Sun et al. 1989, 1991; White and Haber 1990; Sugawara and Haber 1992) were proposed to account for DNA damage-induced mitotic and meiotic gene conversion (reviewed in Thaler and Stahl 1988; Bel- maaza and Chartrand 1994). Gap repair models propose that 3′ ends of broken DNA invade an undamaged homol- ogous duplex, producing two Holliday junctions, and prime DNA synthesis using the undamaged duplex as a template to fill the gap. Gap repair models suggest that most conversion occurs in a gap (i.e., broken alleles are preferentially converted), but allow conversion by repair of hDNA adjacent to gaps formed during strand invasion or branch migration of Holliday junctions. In either type of model, Holliday junctions may be resolved in two senses leading to reciprocal exchange of flanking markers in a half of the conversions. Repair of mismatched bases in recombination interme- diates (hDNA repair) is thought to mediate most or all meiotic gene conversion in yeast (reviewed by Petes et al. 1991). Mismatch repair has been studied extensively in procaryotes and eucaryotes (reviewed in Grilley et al. 1990; Modrich 1991). Escherichia coli has a long patch re- pair system, which excises a single strand between mis- matched bases and a hemimethylated GATC sequence up to several kbp away (Lahue et al. 1989), and short patch repair systems that recognize specific mismatches (Lu and Chang 1988; Radicella et al. 1988; Lieb 1991). Long mis- match repair tracts (> 900 bp) have been observed in yeast (Bishop and Kolodner 1986; Detloff and Petes 1992), and Curr Genet (1996) 29: 335 – 343 © Springer-Verlag 1996 Received: 7 August 1995 / 19 September 1995 Yi-shin Weng · Jennifer Whelden · Laura Gunn Jac A. Nickoloff Double-strand break-induced mitotic gene conversion: examination of tract polarity and products of multiple recombinational repair events ORIGINAL PAPER Y.-s. Weng · J. Whelden · L. Gunn · J. A. Nickoloff () Department of Cancer Biology, Harvard University School of Public Health, 665 Huntington Avenue, Boston, MA 02115, USA Communicated by R. Rothstein