The atpF group-II intron-containing gene from spinach chloroplasts is not spliced in transgenic Chlamydomonas...

In order to determine whether the group-II trans-splicing machinery of the chloroplast of Chlamydomonas reinhardtii can splice a heterologous group-II cis intron, the atpF gene of spinach was transferred into the chloroplast genome of C. reinhardtii using the atpX expression vector. The atpF gene contains a group-II intron which, like other higher plant chloroplast introns, does not self-splice in vitro. The chimeric transgene was expressed at high levels, based on the accumulation of the precursor; however, spliced products could not be detected by Northern blotting, or by RT-PCR coupled with Southern-blot hybridization of the amplified products with an exon-junction probe. These results indicate that the spinach atpF intron is not spliced in transgenic C. reinhardtii chloroplasts. Thus, splicing of chloroplast introns mediated by cellular factors may be species-specific; alternately, the group-II splicing machinery of C. reinhardtii is specific for trans spliced introns.