Homeologous recombination in plastids

In E. coli, sequence divergence dramatically reduces the rate of homologous recombination. A 10% reduction in identity between DNA sequences reduces homologous recombination frequency by 40-fold (Shen and Huang 1986). The effect of mismatches on integration of transgenes can be studied by using heterologous plastid DNA to target integration of foreign genes into plastids. N. tabacum plastid DNA has been used to target integration of foreign genes in Lycopersicon esculen-tum (tomato; Ruf et al. 2001), Petunia hybrida (Zubko et al. 2004) and Solanum tuberosum (potato; Sidorov et al. 1999). Because the transformation frequency of homeologous N. tabacum plastid DNA was not compared with homologous plas-tid DNA the influence of mismatch on plastid transformation frequency in these species is not known. In a more detailed study, a 7.8 kbp region of Solanum nigrum plastid DNA was introduced into N. tabacum plastids (Kavanagh et al. 1999). Recombinant plastid genomes exhibited a mosaic structure comprised of several patches of S. nigrum DNA interspersed with N. tabacum plastid DNA sequences (Fig. 10b). This is consistent with multiple recombination events during integration of 7.8 kbp S. nigrum plastid DNA and random resolution of Holliday junctions. Although S. nigrum and N. tabacum plastid DNA showed 2.4% sequence divergence, plastid transformation frequencies were not reduced relative to using homologous sequences for N. tabacum plastid transformation. Stringent mismatch repair processes which reduce recombination between diverged DNA sequences (Evans and Alani 2000) might be suppressed in flowering plant plastids (Kavanagh et al. 1999). In contrast, homeologous plastid DNA exhibiting around

Fig. 12. Expansion and deletion of short direct repeats by replication slippage (Lovett 2004). Slippage in the daughter strand increases the number of repeat units while slippage in the template strand reduces the number of repeat units.

2-4% mismatch decreased transformation frequency by two to fivefold (Newman et al. 1990) in C. reinhardtii. This might indicate a more efficient plastid mismatch repair system in this green alga.

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