Abstract
In the parasexual hybrid callus between Nicotiana glauca and N. langsdorfii, the two parental types of chloroplasts separate during callus development. The system to investigate the mode of separation involves the hormone independent growth of the hybrid callus between N. glauca and N. langsdorfii as a selective marker for the parasexal hybrid and the isoelectrofocusing patterns of the large subunit (LS) of Fraction I protein as a chloroplast genetic marker. We improved this system by utilizing N. gossei as a chloroplast donor, because the isoelectrofocusing patterns of LS from N. gossei and that from N. langsdorfii can be distinguished clearly. The hormone independency which is suitable for selecting hybrids in the state of small calli was restored by transmitting the nuclear genome of N. glauca into N. gossei by means of sexual hybridization. Using the improved system, the mode of chloroplast separation was investigated in more detail. It was found that 84% of the cells in a two month old parasexual hybrid callus have only one or the other parental type of chloroplasts and 16% of the cells have two types.
We calculated the rate of separation using the formulae based on the assumption that the two types of chloroplasts separate at random by cell divisions in the callus. The result obtained by the calculation turned out to be consistent to the experimental data obtained by the analysis of the two month old parasexual hybrid callus, showing that our assumption is reasonable.
